Assessment of Birth Outcomes in Eswatini After Transition to Dolutegravir-based Treatment
Conditions: Neural Tube Defects, Congenital Abnormalities, Teratogens
Sponsor: Elizabeth Glaser Pediatric AIDS Foundation

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 1 of 54 
 
 
 
 
 
 
ASSESSMENT OF BIRTH OUTCOMES IN ESWATINI AFTER 
TRANSITION TO DOLUTEGRAVIR-BASED TREATMENT 
 
 
Principal Investigators:  
Caspian Chouraya, MD, MSc; Michelle Gill, PhD, MPH; Bonisile Nhlabatsi, MPH 
 
 
Institutions:  
Ministry of Health, Mbabane, Eswatini; Elizabeth Glaser Pediatric AIDS Foundation (EGPAF), 
Mbabane, Eswatini and Washington, DC, USA; George Washington University, Washington, DC, USA 
 
Protocol Version 3.0, dated 26 August 2022 
 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 2 of 54 
Study Team 
 
Name, Surname and Contact Details 
Roles and Responsibilities 
Co-Principal Investigators 
Dr. Caspian Chouraya 
Country Director  
EGPAF Eswatini  
MVA Fund Office Park 
Mbabane, Eswatini 
Email: cchouraya@pedaids.org  
Phone: +268 24041811 
Responsible for the in-country implementation 
of the evaluation, including staff recruitment, 
monitoring study progress, ensuring adherence 
to the scientific goal and ethical standards and 
facilitating bidirectional communication with 
evaluation team; ensuring transparent and 
responsible program and financial management 
of the evaluation and overseeing data 
collection, analysis, report writing, and 
dissemination. Contributes to protocol 
development, overall conduct and management 
and the scientific integrity of the study at 
country level. Ensures government and 
stakeholder involvement and coordination.  
Dr. Michelle Gill 
Associate Research Director  
EGPAF US 
1140 Connecticut Ave NW, Suite 200 
Washington, DC 20036 USA 
Email: mgill@pedaids.org 
Phone: +1 2024488403 
Responsible for protocol development, overall 
conduct (including ethical), management, and 
the scientific integrity of the study at global 
level. Monitors study progress to ensure study 
objectives will be met. Leads interpretation of 
study findings, manuscript writing and 
dissemination and results utilization. Liaises 
with in-country PI and other investigators and 
the donor. 
MOH Principal Investigator 
Bonisile Nhlabatsi 
Program Manager 
Ministry of Health, Eswatini 
Sexual Reproductive Health Unit (SRHU) 
Email: thandazabn@yahoo.com  
Phone: +268 76114263/ +268 24050003 
Contributes to the implementation of the study, 
guiding protocol development and study 
implementation to ensure alignment with 
national policies and priorities and ensure 
transparent bidirectional communication with 
relevant units within the MOH at regional and 
national levels. 
Co-Investigators 
Dr. Lynne Mofenson 
Senior HIV Technical Advisor  
EGPAF US 
Email: lmofenson@pedaids.org 
Phone: +1 2024077970 
 
Provides support to the study team, both global 
and in-country. Will provide technical 
leadership in the implementation of research 
and evaluation projects, and contribute to 
results interpretation and write-up, development 
of publications, and results utilization.    

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 3 of 54 
Dr. Vincent Tukei  
Research Director  
EGPAF Eswatini 
Email: vtukei@pedaids.org  
Phone: +268 78086511 
 
Advises on clinical study issues and 
interpretation of clinical data. Liaises with 
EGPAF and MOH on program and study 
related issues. Will help to ensure that 
experiences from the study are used to inform 
the ongoing national surveillance system and 
government and stakeholder involvement and 
coordination. 
Philisiwe Khumalo 
Senior Public Health Evaluation Manager 
EGPAF Eswatini  
Email: pkhumalo@pedaids.org 
Phone: +268 24048081 
 
Contributes to the day-to-day implementation 
of study activities, including translation of 
evaluation tools and SOPs, training and 
supervision, ethics and regulatory compliance, 
management of data abstraction/collection 
activities, quality, and analysis and 
dissemination and serve as the liaison for 
communication and coordination with ethical 
review boards at the Ministry of Health.  
Dr. Heather Hoffman 
The George Washington University  
Milken Institute School of Public Health 
Biostatistics and Bioinformatics Department 
Email: hhoffman@gwu.edu  
Phone: +1 2029948587 
Responsible for developing statistical analysis 
plan, lead analysis of data, and contribute to the 
write up of study findings. 
MOH Co-investigators 
 
Ntombikayise Dlamini-Madlopha 
Maternal and Newborn Officer 
Ministry of Health, Eswatini 
Sexual Reproductive Health Unit (SRHU) 
Emial: kayisedee@gmail.com   
Phone: +268 76086890 
Contribute to the design and implementation of 
the study, guide protocol development and 
study implementation to ensure alignment with 
national policies and priorities and ensure 
transparent bidirectional communication at 
health facility level. Also contribute to the 
overall management of data 
abstraction/collection activities, data quality, 
and analysis and dissemination activities. 
Wiseman Mngometulu 
PMTCT Focal, Community Mentor Mother 
Coordinator 
Ministry of Health, Eswatini 
Sexual Reproductive Health Unit (SRHU)  
Email: wisdomwisey2@gmail.com 
Phone: +268 76187748 
Contribute to the design and implementation of 
the study, guide the development and 
implementation to ensure alignment with 
national policies and priorities and ensure 
transparent bidirectional communication at 
health facility level. Also contribute to data 
quality assurance, review and interpretation 
and dissemination activities. 
Nompumelelo Mthunzi 
Senior Monitoring and Evaluation Analyst 
Contribute to the design and implementation of 
the study, guide the development and 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 4 of 54 
Ministry of Health, Eswatini 
Strategic Information Department (SID) 
Email: mpumiemthunzi@gmail.com 
Phone: +268 24047252, +268 76210119 
implementation to ensure alignment with 
national policies and priorities and ensure 
transparent bidirectional communication at 
health facility level. Also contribute to data 
quality assurance, review and interpretation 
and dissemination activities. 
 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 5 of 54 
Table of Contents 
Acronyms ...................................................................................................................................................... 6 
Protocol Summary ........................................................................................................................................ 7 
Background & Scientific Rationale .............................................................................................................. 9 
Study Objectives ......................................................................................................................................... 12 
Study Design & Methods ............................................................................................................................ 12 
Study Design Summary .......................................................................................................................... 12 
Site Selection .......................................................................................................................................... 13 
Study Population ..................................................................................................................................... 14 
Sample Size Calculation ......................................................................................................................... 14 
Study Procedures .................................................................................................................................... 15 
Enrolment: ........................................................................................................................................... 15 
Chart abstraction: ................................................................................................................................ 16 
Interviews with women ....................................................................................................................... 17 
Training ............................................................................................................................................... 18 
Study Variables and Definitions ............................................................................................................. 19 
Data Management ....................................................................................................................................... 21 
Confidentiality Protections ..................................................................................................................... 22 
Statistical Analysis Plan .............................................................................................................................. 23 
Ethical Considerations ................................................................................................................................ 24 
Informed Consent Procedures ................................................................................................................. 24 
Risks and Benefits................................................................................................................................... 25 
Request for Consent Waiver ................................................................................................................... 25 
Serious Adverse Events Reporting ......................................................................................................... 26 
Compensation for Participation .............................................................................................................. 26 
Ethical Review ........................................................................................................................................ 26 
Limitations .................................................................................................................................................. 26 
Dissemination and Utilization ..................................................................................................................... 26 
Estimated Study Timeline ........................................................................................................................... 27 
References ................................................................................................................................................... 28 
 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 6 of 54 
Acronyms 
 
ANC  
Antenatal care  
APMR  
ART Patient Monitoring and Reporting 
APR  
Antiretroviral Pregnancy Registry 
ART  
Antiretroviral therapy  
ARV  
Antiretroviral 
CMIS  
Client management information system  
DTG  
Dolutegravir 
EFV 
 
Efavirenz 
EGPAF 
Elizabeth Glaser Pediatric AIDS Foundation 
EHHRRB 
Eswatini Health and Human Research Review Board 
IPD 
 
Inpatient Department 
IRB 
 
Institutional Review Board 
LMP  
Last menstrual period  
MOH  
Ministry of Health 
NNRTI 
Non-nucleoside reverse-transcriptase inhibitor 
NRTI  
Nucleoside reverse transcriptase 
NTD  
Neural tube defect 
OPD  
Outpatient Department 
PI 
 
Protease inhibitor 
PIN 
 
Personal identity number 
PMTCT 
Prevention of Mother-to-Child Transmission 
PrEP   
Pre-exposure prophylaxis 
SID 
 
Strategic Information Department 
SRHU  
Sexual Reproductive Health Unit 
WHO  
World Health Organization  
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 7 of 54 
Protocol Summary  
 
Dolutegravir (DTG) is a second-generation integrase inhibitor that has demonstrated improved 
virologic efficacy and tolerance compared to efavirenz (EFV)-based first-line ART.  Initial data 
in May 2018 from the Tsepamo study in Botswana had suggested a potential ~9-fold increase in 
risk of neural tube defects (NTD) among infants delivered to women receiving DTG at the time 
they became pregnant.  However, in the most recent analysis through March 2022, NTD 
prevalence had stabilized at a low level, 0.11% (10 NTD in 9,460 exposures to DTG at 
conception); this is now the same as the NTD prevalence of 0.11% with non-DTG ART at 
conception (25 NTD in /23,664 births) and is compared to 0.07% in women without HIV infection 
(108 NTD in 170,723 births).  Based on previous data from this study in 2020, demonstrating an 
earlier decrease in NTD prevalence, and updated risk-benefit analyses demonstrating that the 
benefits of DTG significantly outweigh the potential risks, the World Health Organization 
recommends DTG as a preferred first-line drug for treatment, but also indicates continued 
surveillance is needed to more definitively confirm or refute the NTD signal among women of 
reproductive potential.  
 
To contribute to the evidence base on birth defects among women on DTG at conception and to 
support the national birth surveillance system that can be used to determine the rate of birth defects 
in the general population and evaluate outcomes of emergent therapies, we will be conducting an 
observational study at five high volume hospitals in Eswatini, in which birth defect and other data 
will be collected at the time of delivery for all women. The overall aim of this study is to evaluate 
the birth outcomes of HIV-positive women who are receiving DTG or other ARV drug regimens. 
The primary research objectives are to: 1) determine the proportion of neural tube defects among 
live and stillborn infants delivered by HIV-positive women on DTG at conception; 2) determine 
the proportion of neural tube defects among live and stillborn infants delivered by HIV-negative 
women; and 3) determine the proportion of neural tube defects among live and stillborn infants 
delivered by HIV-positive women on non-DTG ART at conception.  
 
Data collection over an approximate 24-month period will involve two main components. The first 
is birth surveillance designed to capture the deliveries of all women presenting to study hospitals, 
including HIV-positive women with DTG exposure both before and during pregnancy or on 
another or no ART regimen and HIV-negative women as well as any stillbirths. Mothers’ 
pregnancy history, birth outcomes, and HIV and ART information (as applicable) will be collected 
from existing clinic registers, reporting forms and databases. Secondly, more detailed information 
on health history, risk factors, and descriptions of birth defects and classification by clinical staff, 
will be collected through interviews and photographs with consenting mothers whose children 
have been identified by clinical staff to have major external birth defects at the time of delivery, 
or in the event of a medical abortion following the identification of a serious fetal birth defect 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 8 of 54 
through ultrasonography. This information will be sent to an external pediatric specialist for a final 
confirmatory diagnosis.  
 
NTD will be the primary study outcome. Key secondary outcome variables include other major 
and surface birth defects and other pregnancy outcomes that could be associated with ARV 
exposure, such as preterm delivery, low birthweight and stillbirths. The primary analysis will be 
descriptive, to determine the proportion of NTD among infants delivered by HIV-positive women 
on DTG at conception and HIV-negative women.  Exploratory statistical investigations may be 
performed for birth defects or other outcomes among women living with HIV receiving DTG at 
conception, women living with HIV receiving non-DTG ART at conception, and women without 
HIV infection.    If assumptions are met for NTD prevalence and the expected number of deliveries, 
one year of surveillance from these hospitals could provide the sample size needed to rule-out a 
relatively rare event like an NTD: approximately 10,000 women on DTG at conception.  
 
Results will be disseminated by EGPAF to the Eswatini MOH at regional and national levels and 
other key stakeholders in-country as well as to a global audience to share information critical to our 
understanding of birth defects and other birth outcomes, among women on DTG at conception and 
in the general population.  
 
 
 
 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 9 of 54 
Background & Scientific Rationale  
 
Ensuring people living with HIV are on optimal ART regimens is a critical component to 
reaching the UNAIDS 95-95-95 goals by 2030 [1]. Dolutegravir (DTG) is a second-generation 
integrase inhibitor that has demonstrated improved virologic efficacy and tolerance compared to 
efavirenz (EFV)-based first-line ART [2, 3]. DTG has a high barrier to resistance, and in the face 
of growing transmitted and acquired drug resistance, especially to non-nucleoside reverse-
transcriptase inhibitors (NNRTIs) like EFV, DTG offers strong first-line therapy [4, 5]. DTG 
has also been shown to be effective and safe when used for second-line therapy in combination 
with an optimized nucleoside reverse transcriptase (NRTI) backbone in individuals with 
virologic failure on NNRTI or protease inhibitor (PI)-based first-line regimens [6-8].  Two 
randomized clinical trials in pregnant women starting ART after the first trimester have 
demonstrated that DTG ART more rapidly decreased viral load than EFV ART, with a 
significantly larger proportion of women having undetectable viral load at delivery with DTG 
than EFV ART [9, 10].   
 
However, in May 2018, investigators from the Tsepamo study, a multi-site surveillance of birth 
outcomes for HIV-positive and HIV-negative women at hospitals in Botswana reported a potential 
signal for neural tube defects (NTD) with use of DTG ART at conception [11].  Four NTD were 
identified in 426 (0.94%) women receiving DTG at the time they became pregnant. This was 
significantly greater than all other comparison groups, including women on non-DTG ART 
regimens at the time of conception, 14 NTD/11,300 deliveries, prevalence 0.12%; starting DTG 
during pregnancy, 0 NTD/2,812 deliveries; and HIV-negative women, 61 NTD/66,057 deliveries, 
prevalence 0.09%.  The study expanded from eight to 18 sites, covering 72% of all births in 
Botswana.  Repeat analysis of data through March 2019 demonstrated a decrease in NTD 
prevalence with receipt of DTG at conception to 0.30% (five NTD among 1,683 deliveries) [12].  
While the prevalence of NTD declined from the initial report, the NTD prevalence remained 
significantly higher than observed with non-DTG ART at conception (0.10%), EFV at conception 
(0.04%), or DTG started during pregnancy (0.04%), or in HIV-uninfected women (0.08%). 
Additionally, the Botswana Ministry of Health and Wellness expanded surveillance of birth 
outcomes at 22 selected non-Tsepamo health facilities, expanding coverage to 92% of all births in 
Botswana when combined with Tsepamo sites [13]. From October 2018 through March 2019, at 
the MOH sites, one NTD was identified among 152 women receiving DTG at conception 
(prevalence 0.66%) compared to no NTD among 381 HIV-positive women receiving non-DTG 
ART at conception and two among 2,328 HIV-uninfected women (0.09%).    
 
A more recent analysis of the Tsepamo study including data through April 2020 was presented at 
the International AIDS Conference in July 2020.  The prevalence of NTD with DTG at conception 
further decreased to 0.19% (seven NTD/3,591 deliveries), remaining stable at this rate from 
September 2019 through April 2020 [14].  In comparison, NTD prevalence with non-DTG ART 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 10 of 54 
at conception was 0.11% (21 NTD/19,361 deliveries), with the prevalence difference between 
DTG vs non-DTG at conception no longer statistically significantly different (prevalence 
difference 0.09%, 95% CI -0.03, 0.30%).  The prevalence difference with DTG at conception did 
remain significantly higher than EFV at conception (eight NTD/10,958 deliveries, 0.07%, 
prevalence difference 0.12%), DTG started during pregnancy (two NTD/4,581 deliveries, 0.04%, 
prevalence difference 0.15%), and HIV-uninfected women (87 NTD/119,630 deliveries, 0.07%, 
prevalence difference 0.12%). Finally, the most recent results from the Tsepamo study, including 
data through March 2022 was presented at the International AIDS Conference in July 2022.  NTD 
prevalence had stabilized at a low level, 0.11% (10 NTD in 9,460 exposures to DTG at 
conception); this is now the same as the NTD prevalence of 0.11% with non-DTG ART at 
conception (25 NTD in /23,664 births) and is compared to 0.07% in women without HIV infection 
(108 NTD in 170,723 births).  The prevalence of NTD in infants born to women on DTG at 
conception does not substantially differ from other exposure groups; prevalence difference 
between DTG versus non-DTG at conception (0.00%, 95% CI -0.07, 0.10) and versus HIV-
uninfected women (0.04%, 95% CI -0.01, 0.13).1  
 
Outside of the Botswana studies, one other NTD has been reported in the January 2020 
Antiretroviral Pregnancy Registry (APR) out of 382 women receiving DTG at the time of 
conception (prevalence 0.26%) [15]. However, most reports in the APR come from higher income 
countries, most with national food folate fortification, which has been shown to be associated with 
a decline in population prevalence of NTD [16, 17]. In Brazil, a retrospective chart review of all 
women with pregnancies and possible DTG exposure included in the national antiretroviral 
therapy database was conducted; the results found no NTDs during the study period in women 
conceiving on either DTG (n=382) or EFV (n=1,045) as a comparator [18]. However, two NTD 
were reported post-hoc in women conceiving on DTG, bringing the estimated NTD prevalence 
with DTG at conception to 0.18% [18, 19]. Brazil has national food folate fortification, and this 
NTD prevalence with DTG at conception is higher than the population NTD prevalence estimate 
of 0.06% [20].  
 
Thus, the available data suggest that if DTG at conception is associated with an increased risk of 
NTD, this risk is likely a ~2 to 3-fold increase over the overall population rates, and not the ~9-
fold increase suggested by the initial Tsepamo data in May 2018. To provide perspective on the 
prevalence increase observed with non-genetic factors associated with an increased risk of NTD, 
maternal diabetes is associated with a 2 to 10-fold increase in NTD, maternal obesity a 1.5 to 3.5-
fold increase, maternal hyperthermia a 2-fold increase, low maternal red blood cell folate levels a 
4-fold increase, and receipt of valproate around the time of conception a 10-fold increase [21, 22]. 
 
 
1Zash R et al. Update on neural tube defects with antiretroviral exposure in the Tsepamo study, Botswana. 24th 
International AIDS Conference, Montreal, abstract PELBB02, 2022. 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 11 of 54 
Based on the decreased NTD prevalence in the updated Tsepamo study and updated risk-benefit 
analyses demonstrating that the benefits of DTG significantly outweigh the potential risks [5, 23, 
24], the World Health Organization (WHO) recommends DTG as a preferred first-line drug for 
treatment as well as a preferred second-line drug in combination with an optimized NRTI 
backbone in individuals failing NNRTI or protease inhibitor –based therapy [25]. The 2019 
guidelines note that women should be provided with information about benefits and risks to make 
an informed choice regarding the use of DTG or other ART, and that continued surveillance is 
needed to more definitively confirm or refute the NTD signal.   
 
Eswatini is an optimal context in which to conduct birth surveillance and generate additional data 
in sub-Saharan Africa in a country without folate food fortification. The adult HIV prevalence is 
one of the highest in the world at 27% and is 35% among pregnant women [26, 27].  The facility 
delivery rate is 87.7% [28]. This relatively small country has 12 public hospitals, allowing for 
focused efforts in facility-level capacity building and data system strengthening in a subset of high 
volume hospitals to allow for robust data collection of birth outcomes in the country. Eswatini 
began transition of HIV-positive adolescents and adults, including women of reproductive 
potential, to DTG-based first- and second-line ART in April 2019. DTG introduction was gradual, 
particularly among women in this group, in an effort to first use existing EFV stock. However, 
transition is expected to ramp up in 2021 and thus, preconception DTG exposures will occur during 
the next couple years in Eswatini, making it an ideal country to institute further birth outcome 
surveillance to provide additional reliable data from Africa to refute or confirm the existing NTD 
association. 
 
EGPAF has been supporting the Kingdom of Eswatini since 2003 to prevent mother-to-child HIV 
transmission (PMTCT) and to provide HIV and AIDS care and treatment services to HIV-positive 
children and adults. Currently, EGPAF supports 65 health facilities in Hhohho and Shiselweni 
regions of Eswatini with PMTCT, HIV prevention (voluntary medical male circumcision, pre-
exposure prophylaxis [PrEP]), HIV testing and counseling, ART and tuberculosis services. 
Together with the Ministry of Health (MOH) and other implementing partners, EGPAF has been 
leading efforts to strengthen birth outcomes monitoring, which will help ensure this surveillance 
study is well integrated into national health programming. 
 
With planned broad transition of programs to DTG-based ART for first- and second-line treatment, 
it will be important to have in place sentinel surveillance to evaluate birth outcomes in women 
who become pregnant while receiving DTG or any new drug regimen. Through this effort, EGPAF 
will enhance the existing surveillance program through sentinel surveillance conducted as part of 
the study in five high-volume public hospitals. The system will allow for the collection of data on 
preconception DTG exposures and birth defects by capturing deliveries from all HIV-positive and 
HIV-negative women, primarily using existing facility sources. This surveillance will strengthen 
and improve upon efforts by the Eswatini MOH to introduce national routine pregnancy outcome 
and infant monitoring, helping to ensure the sustainability of such a system to evaluate new drugs 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 12 of 54 
and interventions as they come to market, such as long-acting antiretrovirals for HIV treatment 
and prevention. Furthermore, there are currently no population-based estimates of birth defects in 
Eswatini. In addition to its utility in providing the comparative group for evaluating the effects of 
DTG exposure, these data will be useful for the country as a whole in describing the general burden 
of birth defects. 
 
Study Objectives  
 
The overall aim of this study is to evaluate the birth outcomes of HIV-positive women who are 
receiving DTG or other ARV drug regimens. The following primary research objectives will be 
addressed in selected hospitals in Eswatini:  
 
1. To determine the proportion of neural tube defects among live and stillborn infants delivered 
by HIV-positive women on DTG at conception. 
 
2. To determine the proportion of neural tube defects among live and stillborn infants delivered 
by HIV-negative women.  
 
3. To determine the proportion of neural tube defects among live and stillborn infants delivered 
by HIV-positive women on non-DTG ART at conception. 
 
 
The secondary research objectives are described below.  
 
1. To determine the general prevalence of major external birth defects among all live and 
stillborn infants regardless of maternal HIV or ART status.  
 
2. To describe pregnancy outcomes (e.g., all major and surface birth defects, preterm delivery, 
low birthweight, miscarriages) among women by HIV status, by ART regimen, by timing 
of exposure to DTG (pre- or post-conception), and other factors such as maternal age at 
delivery and gravida.  
 
Study Design & Methods 
 
Study Design Summary 
This is an observational study at sentinel sites to evaluate the birth outcomes of women receiving 
the standard of care for ART treatment and the outcomes of HIV-negative women, with data 
collected at one time point for all women, at the time of delivery. Collecting data on all women 
who deliver at these sentinel sites with detailed information on any newborns with identifiable 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 13 of 54 
birth defects will be critical to our understanding of birth outcomes and any birth defects. This will 
be achieved by conducting birth outcomes surveillance, with data collection planned on all 
deliveries during an approximate 24-month period. The target population includes all HIV-positive 
women with DTG exposure both before and during pregnancy, HIV-positive women on another 
or no ART regimen, and HIV-negative women delivering at selected hospitals. Detailed 
information will be collected through chart abstraction for all women, and through interviews and 
photographs with consenting mothers whose children have been identified to have major external 
birth defects. 
 
NTD will be the primary study outcome. Key secondary outcome variables include other major 
and surface birth defects and other pregnancy outcomes that could be associated with ARV 
exposure, such as preterm delivery, low birthweight and stillbirths. The primary analysis will be 
descriptive, to determine the proportion of NTD among infants delivered by HIV-positive women 
on DTG at conception, HIV-negative women, and any available women not on DTG. Exploratory 
statistical investigations and modeling may be performed for birth defects or other outcomes among 
the groups described in secondary objective #2 provided there are sufficient sub-group numbers to 
do so. This will also depend on the extent of DTG transition at the time of data collection, 
particularly for comparisons of HIV-positive women on non-DTG regimens and those initiating 
DTG during pregnancy.  
 
Site Selection  
The study will be conducted in high-volume public hospitals (Table 1) across all four regions of 
Eswatini. Five out of the 12 hospitals in the country with the highest number of deliveries in 2020 
were selected purposively. In January 2020 to December 2020, there were 20,501 deliveries in the 
selected hospitals and 6,393 deliveries among HIV-positive women who were on ART in antenatal 
care (ANC) [29].  Of the HIV-positive women on ART in ANC, 85%  were on ART at the first 
visit, resulting in an estimated 5,434 women on ART at the time of conception in these five 
hospitals for the one-year period [26]. Assuming uptake of DTG will range from 65% to 90% for 
the women enrolled during the study period and that the delivery numbers for HIV positive women 
will be similar to previous programmatic numbers, this is estimated to yield 3,145 deliveries of 
women on DTG at conception. If assumptions are met or exceeded for NTD prevalence and the 
expected number of deliveries after one year of surveillance from these hospitals, approximately 
two years of enrollment could provide the sample size needed to rule-out a relatively rare event 
like an NTD (see Table 2).  
 
Table 1. Study hospitals and number of deliveries, January to December 2020 
 
No 
Region 
Hospital  
# of 
deliveries 
# of HIV 
positive 
women 
# of HIV 
positive 
women 
on ART 
Estimated # of 
women on 
DTG at 
conception* 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 14 of 54 
1 
Hhohho 
Mbabane Government 
Hospital 
5,579 
1,768 
1,767 
872 
2 
Manzini 
Raleigh Fitkin 
Memorial Hospital 
5,534 
1,979 
1,978 
961 
3 
Manzini 
Mankayane 
Government Hospital 
2,516 
7,14 
712 
378 
4 
Lubombo 
Good Shepherd 
Hospital 
3,659 
1,104 
1,097 
531 
5 
Shiselweni 
Hlathikhulu Hospital 
3213 
828 
825 
403 
 
Total 
 
20,501 
6,393 
6,379 
3,145 
*Based on assumptions and previous programmatic estimates 
 
Study Population 
For capture of routine pregnancy information, the study population is defined as all women 
delivering infants at selected hospitals in Eswatini, including women delivering stillborn infants 
and those who would have miscarriages or medical abortions due to identified congenital defects. 
A subset of this population will be recruited for additional data collection. Inclusion criteria for 
this subset are: (1) a birth defect as identified by a midwife or other clinician at the study hospital 
and (2) willing to provide informed consent. For the additional data collection component, women 
of any age will be considered eligible. Adolescents less than 18 years of age, defined as 
emancipated minors due to their pregnancy/motherhood status, will be asked to provide informed 
consent for themselves.  
 
Sample Size Calculation  
The sample size target will be based on updated Tsepamo study findings from 2022, which found 
an NTD prevalence of 0.11% (10/9,460 deliveries) compared to earlier data from 2019 which 
found a prevalence of 0.30% (5/1,683 deliveries) among women receiving DTG ART at 
conception [12].  When the sample size is between 7,500-10,000 for the exposure group of interest, 
a two-sided Wilson score-based 95% confidence interval for a single proportion will have a 
precision of 0.00083-0.00098 for an expected NTD prevalence of 0.11% among deliveries to 
women with DTG at conception exposure with a probability of at least 95% (Table 2)[30].  Sample 
sizes based on the prevalence for the other two groups using the latest Tsepamo study findings are 
also presented (0.07% for HIV-negative women and 0.11% for HIV-positive women not on DTG). 
As we expect the enrollment number for HIV-negative women to be higher than the other groups 
and the number of HIV-positive women on non-DTG regimens at conception to be low as DTG 
transition efforts mature, precision estimates are presented according to these expectations. Larger 
sample sizes will increase the power and decrease the type I error in the statistical models used to 
address secondary objective 2; therefore, we will enroll everyone during the study period. Sample 
sizes are provided in the table below based on varying proportions and precision.  
 
 
Table 2. Sample sizes based on assumed proportions (0.0007, 0.0011) and precision ranges 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 15 of 54 
Proportion 
Precision (Half-Width) 
Sample Size (N) 
Probability 
HIV-negative 
 
0.0007 
0.0013 
3800 
0.946 
0.0014 
3600 
0.957 
0.0015 
3400 
0.966 
0.0016 
3200 
0.973 
HIV-positive 
on DTG at 
conception 
  
0.0011 
0.00083 
10000 
0.968 
0.00085 
9500 
0.962 
0.00087 
9000 
0.955 
0.00090 
8500 
0.946 
0.00095 
8000 
0.965 
0.00098 
7500 
0.958 
HIV-positive 
on non-DTG 
at conception 
   
0.0011 
0.0029 
1500 
0.974 
0.0032 
1400 
0.980 
0.0034 
1300 
0.985 
0.0035 
1200 
0.955 
0.0036 
1100 
0.965 
 
As there is some uncertainty with respect to the number of deliveries that will be observed among 
women receiving DTG ART at conception, enrollment numbers will be reviewed monthly for 
progress towards the study sample size to assess the likely trajectory for achieving the target 
sample size. All deliveries will be captured to help ensure we are positioned to detect an association 
between DTG preconception exposure and NTD, if one exists, and to be able to describe the 
prevalence of NTD in the general population.  
 
Study Procedures 
Enrolment:  
Data collection will involve two main components: 1) birth surveillance designed to capture the 
deliveries of all women presenting to the five study hospitals and 2) interviews with women and 
photographs taken of their newborns with possible defects identified at birth. For each delivery, 
midwives or data collectors will complete a paper-based version of the study enrollment log (or 
master linking document) by writing in the date of admission, date of delivery, unique patient 
identifiers (e.g., Personal Identity Number [PIN], ANC and ART as applicable), ANC health 
facility name and the woman’s name and her phone number, which are necessary to locate and 
link women’s information in the registers and other sources (e.g., CMIS, APMR) and confirm her 
contact information in the case of duplicated/multiple records. To ensure that delivery data are 
captured in real-time (including weekends and overnight), facility-based nurse midwives will 
assist at the first point of data collection, particularly during off-work hours, when feasible. Per 
routine practice, nurse midwives will examine the infant for external birth defects; examinations 
of stillborns will be conducted if feasible. For each woman’s entry on the log, the outcome of this 
examination will be documented, to capture whether or not a major external birth defect was 
identified. A generic term (e.g., ‘determination’) and an accompanying numeric response code will 
be used so those outside the study would not know the meaning of the code, to avoid capturing 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 16 of 54 
any sensitive information in this log. This would then help to determine the next data collection 
step for each woman: recruit, consent, and interview to capture more detailed information on her 
health history and risk factors if there is a birth defect; if not, the data clerk will proceed with data 
abstraction. Additional procedures for each of these two steps are described below.  
 
Chart abstraction: 
Trained data collectors will be stationed at each study hospital and will be primarily responsible 
for data collection of still- and live-born deliveries at maternity, and any miscarriages or medical 
abortions due to identified congenital defects of women presenting to the facilities 
(Gynecology/outpatient department [OPD]/inpatient department [IPD]) to the extent possible. 
Each day, data collectors will review the enrollment logs completed by midwives and cross-
reference those forms with the clinic registers to verify that all deliveries have been captured. They 
will abstract basic information about the mother, child, pregnancy and delivery using registers, 
patient handheld cards, and other paper-based or electronic sources, such as the national client 
management information system (CMIS) and the ART Patient Monitoring and Reporting (APMR), 
to the extent necessary, and enter into a study record using an electronic form loaded onto an 
electronic device. Since routine information will be collected from the existing records, and does 
not include any primary data collection with participants, the activity presents no more than 
minimal risk of harm to participants and  a waiver of consent is requested. 
 
The main study data sources used in hospitals will be the maternity and perinatal registers and the 
ANC card. Data, which are not available in the hospital registers or the ANC card, will be accessed 
in the following way: 
 
• Data from women’s electronic database entries will be extracted and integrated into their 
study records. This will be done by data collectors at hospitals using the onsite databases 
or by study staff accessing them offsite (e.g., at the EGPAF national office). For the latter, 
this is likely to occur when additional data collection support is needed or when the older 
version of an electronic database like CMIS, APMR, or the health management information 
system (HMIS) needs to be accessed, for example, when women received care at an ART 
clinic that has not yet been updated to the current electronic data capture system. Variables 
include PrEP regimen and start and stop dates for women who were HIV negative at the 
time and for women with HIV, ARV drugs taken at current regimen (as applicable) and 
date of last menstrual period (LMP) for women presenting to the gynecological ward or 
other department with miscarriages or for recommended medical abortions and for women 
presenting to maternity if it has not already been captured by midwives.  
 
As part of completion of the enrollment log for women presenting to maternity, midwives or data 
collectors will capture the women’s LMP from her handheld ANC card (if she has brought her 
card and this information is documented in her card), as this is critical to determining primary 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 17 of 54 
outcomes, and other information as needed (see Table 3). Receipt of PrEP in pregnancy will also 
be captured on the log, and a ‘yes’ response will prompt data collectors or the larger study team to 
extract regimen and start and stop dates from electronic sources as needed and as described above 
for the women’s study record.  
 
Information about miscarriages (or spontaneous abortions, defined as < 28 weeks gestation), will 
also be captured to the extent possible, but will be dependent on women’s self-report and 
presentation to the gynecological ward. Similarly, information about medically induced abortions 
will also be collected as feasible (described further below). Data collectors will make regular, 
frequent visits to the ward or other departments co-located at the same hospital as the maternity to 
check for any miscarriages (which may be found in the gynecological inpatient and outpatient 
registers or improvised registers) or medical abortions. Clinic staff may also notify study team 
members of women potentially meeting study criteria outside of the maternity ward.  Similar 
information will be abstracted for women with miscarriages and abortions (to the extent it is 
available) and live or still births.  
 
Interviews with women 
A woman who delivers an infant with an identified major external birth defect will be referred by 
clinic staff and recruited for enrollment into a separate study component. Because women who 
have abortions will only be recruited for the study if the pregnancy was terminated due to the 
presence of a birth defect detected by imaging, attempts will also be made to interview them. If 
this is not feasible, for instance if the woman has already been discharged or the midwife is 
unavailable, her information will be abstracted from existing records, similar to the procedure for 
miscarriages as described above, plus information on the identified congenital defect.  These data 
may be collected retrospectively (while still within the study period) or prospectively. 
 
If she is willing to hear more about the study, she will undergo an informed consent process. Nurses 
and data collectors will be trained in the administration of informed consent, and either may 
perform this task. Women will be interviewed for more in-depth information on her HIV testing 
and ART history, other medical history of selected conditions (e.g., diabetes, malaria, TB) and 
potential teratogenic exposures. Data will be verified from the women’s handheld patient cards 
and other medical records as available. Accompanying photographs of the infant will also be taken 
(with consent) from multiple perspectives to provide a complete picture of the defect.  
 
As routine practice, doctors based at the maternities should examine any infant identified by the 
nurse midwife as having an external defect and consult on the diagnosis. As part of surveillance, 
existing EGPAF mentors and the study team will support efforts to standardize and strengthen this 
practice. The interview responses, which will include relevant details of the major structural 
abnormality through closed and open-ended questions, and photographs (if available), will be sent 
to an external expert/specialist, for a final confirmatory diagnosis by the study coordinator or other 
study team member. This expert will have training in medical genetics or a similar field and have 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 18 of 54 
experience with these types of reviews for other studies or registries. The specialist will review the 
information (first stripped of HIV and ART status) and diagnosis made by the site level clinical 
team.  It will not include the woman’s name, contact information or the hospital name; she will be 
identified by her study identification number only. Healthcare workers will receive feedback on 
the diagnoses made by the specialist. If the diagnosis by the specialist differs from the diagnosis 
made by the clinical staff, healthcare workers in study sites may contact the mother (with 
permission and phone number obtained at the time of the interview) to inform her of the final 
diagnosis and provide any further information or referrals as needed and appropriate.  
 
Below is a figure that summarizes the flow of information for possible birth defects identified at 
surveillance sites, including reporting to the donor (ViiV Healthcare) and to the institutional 
review boards (IRBs).  
 
Figure 1. Flow Chart for Possible Major External Birth Defects Identified at Surveillance Site 
 
 
 
Training 
As part of the study, health providers (doctors, nurses) in each hospital maternity will receive 
training to further strengthen their skills in examination and identification of birth defects. The 
study coordinator will also provide support to midwives at maternities for surveillance activities. 
The study coordinator and investigators will conduct regular monitoring visits to the hospitals to 
provide mentorship on performing examinations to ensure birth defects are accurately classified 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 19 of 54 
and to review surveillance forms against facility records for quality assurance. In addition, 
mentorship and supportive supervision are already provided by implementing partners (including 
EGPAF) and by the MOH on a regular basis. Training and ongoing supportive supervision will be 
provided to nurse midwives and other providers at study hospitals to strengthen documentation 
practices, in addition to skills building for conducting examinations and identifying birth defects.  
 
Study Variables and Definitions  
For all deliveries/births, routine data will be captured on mother’s pregnancy history, birth 
outcomes, and HIV and ART information as applicable. Variables to be collected will include: 
• Mother’s age, gravidity, parity, LMP, and HIV status 
• Pregnancy outcomes: stillbirth (≥ 20 weeks), prematurity (< 37 weeks), delivery mode, 
infant birth weight, any major surface birth defects, including NTD, and neonatal death 
before discharge  
• If HIV-positive: ART regimen at time of conception (ARVs, date initiated) and any 
changes through delivery (ARVs, start/stop dates). 
• If HIV-negative: receipt of PrEP in pregnancy (start/stop dates), PrEP regimen 
 
 
Table 3 below presents the variables to be collected through surveillance on all women and 
probable variable sources. They are subject to change, and will be ultimately determined by what 
is routinely collected under the program, as we are relying on existing data. Tool versions and 
instructions for completion may change over time and we will align our data collection 
accordingly.  
 
Table 3. Illustrative variables to be collected for routine surveillance (all women), by likely source  
 
Variables 
Source(s) 
Pregnancy and 
HIV history  
Mother’s age 
Maternity register, Perinatal register 
ANC facility  
Maternity register, ANC card  
Last menstrual period  
ANC card 
Gravidity 
Maternity register 
Parity 
Maternity register, Perinatal register 
HIV status and date of last test prior 
to L&D  
Maternity register, Perinatal register 
HIV retesting (eligibility, testing date 
and result) 
Maternity register 
HIV testing if not tested previously 
(testing date and result) 
Maternity register  
Gestational age at first ANC 
Perinatal register  
Birth outcomes 
Gestational age at delivery  
Perinatal register 
Mode of delivery 
Maternity register, Perinatal register 
Date of delivery 
Maternity register, Perinatal register 
Stillbirth (FSB, MSB) 
Maternity register, Perinatal register 
Sex of baby 
Maternity register, Perinatal register 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 20 of 54 
Weight at birth  
Maternity register, Perinatal register 
Single or multiple birth  
Free form text in maternity register* 
Neonatal death before discharge: 
cause, baby age at death  
Perinatal register 
ART/PrEP  
Current ART regimen and date 
started 
CMIS, APMR, Perinatal register, handheld 
ART card 
ART regimen at time of conception 
(if different from above), start/stop 
dates 
CMIS, APMR, handheld ART card 
Any other ART regimen taken during 
pregnancy (if applicable), start/stop 
dates 
CMIS, AMR, handheld ART card 
Receipt of PrEP in pregnancy  
ANC card 
Current PrEP use, regimen, start/stop 
dates 
CMIS, ANC card, APMR 
For women presenting to health facility with miscarriages and medically induced abortions**   
Pregnancy and 
HIV history 
Mother’s age 
Gyn OPD/IPD registers, ANC card 
Last menstrual period  
CMIS, ANC card 
Gravidity 
Gyn/OPD/IPD registers, ANC card 
Parity 
Gyn/OPD/IPD registers, ANC card 
Pregnancy condition  
Gyn/OPD/IPD registers, ANC card 
HIV status 
Gyn/OPD/IPD registers, ANC card, CMIS, 
APMR, handheld ART card 
ART/PrEP 
On ART 
Gyn/OPD/IPD registers, ANC card, CMIS, 
APMR, handheld ART card 
Current ART regimen and date 
started 
CMIS, APMR, ANC card, handheld ART 
card 
ART regimen at time of conception 
(if different from above), start/stop 
dates 
CMIS, APMR, ANC card, handheld ART 
card 
Any other ART regimen taken during 
pregnancy (if applicable), start/stop 
dates 
CMIS, APMR, ANC card, handheld ART 
card 
Current PrEP use, regimen start/stop 
dates 
CMIS, APMR, ANC card 
Birth defect 
(abortions only) 
Birth defect and description   
Ultrasound scan report 
*Will capture to the extent that this is appropriately documented.  
**Abortions due to identified congenital defects will only be captured under surveillance if it is not 
possible to conduct an interview 
 
For the primary analysis, outcomes will be defined as follows:  
 
HIV-positive status: women with a documented positive result in the perinatal and/or maternity 
register or other clinical source record.  
 
HIV-negative status: women with the last HIV test result recorded during pregnancy or labor and 
delivery as negative.  

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 21 of 54 
 
DTG at conception: (for women on DTG-based regimens): determined using the date of initiation 
of DTG and date of LMP or gestational age at delivery (if LMP not available). Given the 
uncertainty around LMP and dates of conception, we are defining ART at conception as maternal 
ART that started up to 8 weeks after the calculated LMP date, which would be up to 6 weeks after 
the estimated date of conception. This will help to ensure we include the first 4 weeks of gestation, 
the defined period for neural tube closure.  
 
NTD: definite (confirmed by photograph) or probable (diagnosed on the basis of a description but 
with no photograph) myelomeningocele, meningocele, encephalocele, anencephaly with or 
without craniorachischisis or iniencephaly.  
 
Other birth defects: Using the parameters defined by the Tsepamo study, other abnormalities 
identified by routine surface examination before hospital discharge will be classified as major 
external structural malformations if they are deemed to be significant from a clinical, surgical or 
cosmetic standpoint [12]. Also as in Tsepamo, surface examinations will not include those of the 
inside of the mouth, auscultation of the heart, or testing for inguinal hernias, undescended testes, 
or hip dysplasia. Other diagnostics, such as imaging, may be used to make a birth defect 
determination for women who ultimately undergo a medically induced abortion as a result of a 
birth defect identified in utero. Ultrasounds are occasionally used at study hospitals at the request 
of patients or under certain circumstances, though chromosomal testing, mutation analysis and 
autopsy data will not be used.  
 
Data Management  
 
Data will be entered directly into ODK-X database (from tablet or any other configured electronic 
devices) using existing paper-based and electronic clinical record sources. Data collectors will be 
primarily responsible for birth surveillance while data collectors and nurse midwives will be 
responsible for interviews with women and taking photographs. Both will work together to help 
ensure every woman meeting study eligibility criteria is captured. Everyone responsible for 
collecting data or supervising collection will be trained on protocol specific procedures, research 
ethics and data entry procedures into ODK-X. Branching and skip patterns as well as logic checks 
and reasonable ranges will be built into data collection tools to help assure data quality. In addition, 
data collected for the study may need to be triangulated using multiple clinic sources, patient 
handheld cards, and electronic databases for relevant variables; the study team may request 
providers help to reconcile inconsistencies across sources.   
 
Experienced EGPAF data management staff will oversee data entry and maintain the study 
database. The database will be stored on a secure web-based server using the ODK-X platform, 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 22 of 54 
which has built-in encryption capabilities. All electronic devices used in the study will be password 
protected and access to the ODK-X application will include another level of authentication.  
 
Data will be synchronized with the secure cloud-based server maintained by EGPAF Global, 
ideally on a daily basis or later (e.g., weekly) when internet connectivity is intermittent. For data 
that may need to be extracted from electronic sources at the EGPAF Mbabane office, the data 
manager or other data staff will be able to update entries to the most up-to-date database and make 
other modifications as needed. The study database will be automatically backed up daily on 
OneDrive-based repository. The backups will synchronize automatically at multiple specified 
times during the day. In case of data loss in the primary database in country, these backups will be 
used for data recovery. The in-country data manager will be responsible for the daily operations 
of the database, including verifying that the data are synchronizing properly between point of data 
collection and the database. The global data manager will support the in-country team in terms of 
building additional validation controls as needed, verifying that the backups are synchronizing 
properly on the server and routine maintenance of the server. Access to the EGPAF server will be 
restricted to authorized study personnel. 
 
Confidentiality Protections  
There is a minimal risk of confidentiality breach. Clinic and other ID numbers, names and contact 
numbers will be collected to ensure our ability to link data for a complete study record and carry 
out quality assurance practices. In addition, by virtue of their presence in the clinic and data 
abstraction activities from existing clinic records, data collectors and other study team members 
as part of study monitoring, will view individual health information. However, identifying 
information will not be contained in the same dataset with study-related information from the 
abstraction and interviews; this information will only be identified by a unique study identification 
(ID) number.   
 
At each hospital, the study team will maintain a master enrollment log that permits linking of the 
participant through their ID numbers, name and contact number used for health services delivery, 
to their unique study number. As described above, data collectors and midwives will be responsible 
for completing the log. Unique study ID numbers will be preprinted on the enrollment logs. This 
master log will be kept in a locked cabinet in a locked office with limited access at study sites. 
Data collectors will also enter selected information (PIN, clinic numbers, names, phone numbers, 
and study ID numbers) into an abbreviated electronic version of the log loaded onto the electronic 
device. Only selected members of the study team will have access to the paper and electronic 
master documents based on the need to facilitate data collection. The electronic version will have 
even greater restricted access and will be used primarily for quality assurance and to enter any 
updates to the records that can only be done through accessing national databases from a central 
(office) location; data collectors will only view/access the electronic log for their respective sites. 
In addition, all study team members will sign a data handling and confidentiality agreement and 
complete an online ethics course certification. Data will also be stored in password-protected 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 23 of 54 
systems or devices, with prompts to change passwords on a regular basis. Finally, to the extent 
possible, data abstraction will occur in a private location of the health facility to minimize any risk 
of unauthorized viewing of medical records. 
 
Separate enrollment logs will be maintained for women who participate in interviews and will 
include participant names, the unique study ID number assigned to them and the date consented 
and enrolled into the study. If they agree, the log will also include their phone number to be 
contacted in case additional follow-up information is deemed necessary to provide after the 
specialist’s review. Participant names and contact information will not be on any study documents 
linked to their interview responses nor in the study database. To minimize the risk of someone 
inadvertently hearing women’s responses, the study team will work with hospital managers to 
identify private areas for interviews, such as a separate consultation room.  
 
The enrollment logs will be destroyed after completion of data analysis. Other paper-based study 
documents, namely the signed consent forms for women participating in interviews, will be 
retained for five years after completion of the study. After that time, the documents will be 
destroyed and the in-country PI or designee will document the destruction.  
 
Information provided to the specialist on birth defects will be stripped of any identifiers and shared 
securely with appropriate safeguards in place (e.g., password-protection, secure link instead of 
email). They will have access to this information until the final study analyses are completed, then 
we will request the specialist to document the destruction/deletion of all related study materials. 
Statistical Analysis Plan 
 
Summary statistics will be calculated for all variables of interest to describe the sample. 
Specifically, means and standard deviations or medians and interquartile ranges (depending on the 
distribution of the data) as well as minima and maxima will be calculated for all continuous 
variables. Frequency counts and percentages will be reported for all categorical variables.  
 
Primary Objectives 1, 2, and 3: The proportion of NTD among all live and stillborn infants 
delivered by (1) HIV-positive women on DTG at conception and (2) HIV-negative women will be 
calculated as the number of births with NTD divided by the total number of births for each group 
of women, and (3) HIV-positive women on non-DTG ART at conception. The 95% confidence 
intervals around these proportions will be calculated with the Wilson Score method, which is 
appropriate for small proportions close to zero. The difference in the proportions between the three 
groups will be estimated along with the corresponding 95% confidence interval based on the 
Newcombe method. These analyses will be conducted both with and without NTD identified 
through ultrasound for women who underwent medically induced abortions.  Statistical inference 
will only be made if there is sufficient power to do so.  
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 24 of 54 
Secondary Objective 1: The general prevalence (proportion) of major external birth defects 
among all live and still born infants regardless of maternal HIV or ART status will be calculated 
as the number of births with major external birth defects divided by the total number of births to 
all women. The 95% confidence intervals around the prevalence of these birth defects will be 
calculated with the Wilson Score method. 
  
Secondary Objective 2: Frequencies and proportions of pregnancy outcomes (e.g., all major and 
surface birth defects, preterm delivery, low birthweight) will be calculated along with 95% Wilson 
Score confidence intervals for subgroups of women based on HIV status, ART regimen, timing of 
exposure to DTG (pre- or post-conception), and timing of exposure to any other ART regimen. 
Estimates of differences in proportions among HIV-positive women will be considered as follows: 
(1) Women who started on DTG ART preconception and those who started on non-DTG regimen 
preconception; (2) women who started on DTG preconception and those who started on DTG 
during pregnancy; and (3) women who started on DTG preconception and those who started a non-
DTG regimen during pregnancy. Finally, all HIV-positive women on any ART regimen will be 
compared to HIV-negative women. Differences in these proportions between the subgroups of 
women will be examined with simple (unadjusted) and multiple (adjusted) logistic regression or 
log-binomial models. Relative risk (risk ratios) will be reported along with their corresponding 
95% confidence intervals. If reliable statistical estimates cannot be generated, due to a small 
number of cases, we will use Firth’s penalized maximum likelihood approach. Adjusted models 
will control for factors such as maternal age at delivery and gravida. All potential covariates will 
be pre-defined in a separate data analysis plan.  
 
 
Ethical Considerations 
 
Informed Consent Procedures  
Written informed consent will be obtained by midwives or study staff trained in consenting 
procedures prior to the collection of any primary study data. Before conducting an interview, they 
will explain the study to the participant using the informed consent form in their preferred language 
(siSwati or English). This will include the purpose of the study, the risks and benefits of 
participation and how risks will be mitigated. They will explain that participation is voluntary and 
it is their choice to participate in the study; participants will be assured that they may refuse to 
participate or refuse to answer any questions and it will not affect their receipt of any health 
services. The consent form will be read clearly to participants; literate participants or witnesses of 
illiterate participants will be asked to read along. Participants will have the opportunity to ask any 
questions regarding study procedures.  Once the consent form has been read and the participant 
feels all their questions and/or concerns have been addressed, literate participants will be asked to 
sign and date the consent form in the appropriate signature block. For those who are illiterate, a 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 25 of 54 
thumbprint will be used to indicate consent and a witness will observe the consent process, sign 
and date the form to certify that the participant has been read the consent on that day and 
voluntarily agreed to take part in the study. Participants will be offered a copy of the informed 
consent form, which includes study team’s contact numbers, in case they have any additional 
questions about participation. 
 
Women with deliveries who consent for the study will be asked if they agree to 1) participate in 
an interview, 2) have photographs taken of her child (children for multiple births) to help classify 
the birth defect, and 3) be followed up later if the specialist reviewing her case has a different 
diagnosis than the maternity or other pertinent information to share. However, women may still 
participate in the study if they only agree to #1 or #2; if they agree to one or both of these 
components, they may also still participate in the study, even if they do not consent to #3. 
 
Risks and Benefits 
This observational study has minimal risk, with the primary risk being a breach of confidentiality. 
Ways to address this and the risk of privacy breaches have been described above in a previous 
section. Secondly, women participating in interviews could also feel uncomfortable or upset by 
the questions on their previous health history, use of drugs or other therapies, or their HIV status, 
particularly as they are being questioned after learning of a major birth defect in their child or 
undergoing an abortion. The midwives and other study staff who may be involved in this study 
component will be trained in non-biased interview techniques and undergo role-playing in training 
sessions to hone this practice. Participants will also be assured that they do not have to answer any 
question that they do not feel comfortable responding to, and that they may leave the study at any 
time.  
 
There will be no direct benefits to participants from taking part in this study. However, the 
information provided by participants will add make a valuable contribution to the international and 
national evidence base on birth defects and risk factors, which could help to improve pregnancy 
and birth outcomes in the future for other women and their children.  
 
Request for Consent Waiver  
Other than the primary data collected from women with probable or definite birth defects, 
surveillance data will be available through routine collection of clinical data during the provision 
of medical care.  No additional information will be collected outside of what is routinely recorded 
in patient records during standard medical care. For this component, there will be no direct 
interaction with the patients by the data collectors or other study team members for the purposes 
of the study. Therefore, a waiver of informed consent is being requested for the following reasons 
according to the U.S. Code of Federal Regulations (CFR 46.116(f)): 
1. The research presents no more than minimal risk of harm to subjects,  
2. The waiver or alteration will not adversely affect the rights and welfare of the subjects; and 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 26 of 54 
3. The research could not practicably be carried out without the waiver or alteration (given 
the number of deliveries to captured as necessitated by this study design). 
 
 Serious Adverse Events Reporting 
While they are not anticipated, any serious adverse event related to the study, and which the study 
team is made aware of, will be reported promptly to the IRBs and the donor in accordance with 
their respective procedures. Reporting of any birth defects and stillbirths among women on DTG 
will be reported to the donor per Figure 1 above. This information will be summarized in annual 
reports to the IRBs. Any suspected or confirmed breaches in confidentiality will also be reported 
to the IRBs and the study facility will be notified as appropriate.  
 
Compensation for Participation  
Study participants will not be paid to participate in the study, nor will they be provided with 
incentives.  
 
Ethical Review 
This protocol will be reviewed by the Eswatini Health and Human Research Review Board 
(EHHRRB) and an IRB in the US.  
 
Limitations  
 
As this study relies heavily on routinely collected data, the primary study limitation is possible 
incomplete recording by health providers and inaccuracies in the records. As described above, the 
study team will support nurse midwives and other providers at study hospitals to strengthen 
documentation practices to mitigate this risk. Study staff will perform routine quality assurance 
checks and resolve missing and inaccurate data entries to the extent possible through ongoing data 
cleaning and frequent site monitoring. Secondly, as this study takes place at hospitals only, we will 
not be able to capture the outcomes of home deliveries. However, the facility delivery rate in the 
country is about 88%. Similarly, women who experience miscarriages may not present to the 
clinic, so we will likely underrepresent this number. Finally, during the interviews with women, 
there may be a certain degree of social desirability or recall biases as they try to remember for 
instance the name and timing of a particular medication they took, especially after they have just 
learned of a birth defect in their newborn or fetus. Interviewers will be trained to employ tools 
such as calendars, reminders of memorable events like holidays and use photographs or 
descriptions of certain drugs to help stimulate one’s memory. They will also be trained to ask 
questions and receive responses in a way that does not imply judgement.  
Dissemination and Utilization 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 27 of 54 
The Eswatini MOH will work closely with EGPAF to disseminate results at regional and national 
levels and other key stakeholders in-country through study-specific meetings and via existing 
platforms, such as technical working group and research advisory committee meetings. The MOH 
will be involved throughout the study preparation and implementation stages, through MOH 
investigators (from the Sexual Reproductive Health Unit), Health Research and Innovations Unit, 
and Epidemiology and Disease Control Unit to help ensure that data serve the country’s needs for 
birth surveillance and that results can inform the birth surveillance network for Eswatini going 
forward. At regional and global levels, findings from this study will be shared across other EGPAF 
programs and with the donor. Abstract(s) will be submitted for consideration at national and 
international meetings and manuscript(s) will be submitted to an international peer-reviewed 
journal. 
 
The results will inform the ongoing national surveillance program, through the provision of 
feedback on the quality and completeness of key indicators that could be considered for any 
national tool refinements and the surveillance process itself (e.g., data collection, clinic flow, etc.). 
As relevant, data on DTG-based ART and other regimens will also be shared with the Eswatini 
National AIDS Program to contribute to future discussions on national ART guideline updates for 
women of child-bearing age.  
 
Estimated Study Timeline 
 
Below is an estimated study timeline, starting from the time of receipt of IRB approval. Once 
approval of the protocol package has been obtained from Eswatini and US IRBs, the study duration 
is estimated to be 30 months, with approximately one month to develop standard operating 
procedures, train midwives and other study staff and prepare facilities, 24 months for data collection 
and approximately five months following data collection to clean, analyze and disseminate the final 
data. Interim data analysis, writing and dissemination activities will also be conducted while data 
collection is ongoing.  
 
 
  
 
QUARTER (Q) 
ACTIVITY 
Q1 
Q2 
Q3 
Q4 
Q5 
Q6 
Q7 
Q8 
Q9 
Q10 
Study training and 
site preparation  
X 
 
 
 
 
 
 
 
 
 
Data collection  
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Data cleaning and 
analysis 
 
 
 
 
X 
 
 
 
X 
 
Write-up of findings 
 
 
 
 
X 
X 
 
 
X 
X 
Dissemination 
 
 
 
 
 
X 
 
 
 
X 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 28 of 54 
 
 
References 
 
 
1. 
UNAIDS. On the fast-track to an AIDS free generation. Geneva: UNAIDS; 2016 [cited 2018 April 
21]. Available from: www.unaids.org/sites/default/files/media_asset/GlobalPlan2016_en.pdf. . 
2. 
Walmsley S, Baumgarten A, Berenguer J, Felizarta F, Florence E, Khuong-Josses MA, et al. Brief 
Report: Dolutegravir Plus Abacavir/Lamivudine for the Treatment of HIV-1 Infection in Antiretroviral 
Therapy-Naive Patients: Week 96 and Week 144 Results From the SINGLE Randomized Clinical Trial. J 
Acquir Immune Defic Syndr. 2015;70(5):515-9.  
3. 
Venter WDF, Sokhela S, Simmons B, Moorhouse M, Fairlie L, Mashabane N, et al. Dolutegravir 
with emtricitabine and tenofovir alafenamide or tenofovir disoproxil fumarate versus efavirenz, 
emtricitabine, and tenofovir disoproxil fumarate for initial treatment of HIV-1 infection (ADVANCE): 
week 96 results from a randomised, phase 3, non-inferiority trial. The lancet HIV. 2020;7(10):e666-e76.  
4. 
Vitoria M, Hill A, Ford N, Doherty M, Clayden P, Venter F, et al. The transition to dolutegravir 
and other new antiretrovirals in low-income and middle-income countries: what are the issues? Aids. 
2018;32(12):1551-61.  
5. 
Phillips AN, Bansi-Matharu L, Venter F, Havlir D, Pozniak A, Kuritzkes DR, et al. Updated 
assessment of risks and benefits of dolutegravir versus efavirenz in new antiretroviral treatment initiators 
in sub-Saharan Africa: modelling to inform treatment guidelines. The lancet HIV. 2020;7(3):e193-e200.  
6. 
Hamers RL. Dolutegravir for second-line antiretroviral therapy. The Lancet Infectious diseases. 
2019;19(3):218-9.  
7. 
Aboud M, Kaplan R, Lombaard J, Zhang F, Hidalgo JA, Mamedova E, et al. Dolutegravir versus 
ritonavir-boosted lopinavir both with dual nucleoside reverse transcriptase inhibitor therapy in adults with 
HIV-1 infection in whom first-line therapy has failed (DAWNING): an open-label, non-inferiority, phase 
3b trial. The Lancet Infectious diseases. 2019;19(3):253-64. 
8. 
Waitt C, Orrell C, Walimbwa S, Singh Y, Kintu K, Simmons B, et al. Safety and pharmacokinetics 
of dolutegravir in pregnant mothers with HIV infection and their neonates: A randomised trial (DolPHIN-
1 study). PLoS medicine. 2019;16(9):e1002895.  
9. 
Kintu K, Malaba TR, Nakibuka J, Papamichael C, Colbers A, Byrne K, et al. Dolutegravir versus 
efavirenz in women starting HIV therapy in late pregnancy (DolPHIN-2): an open-label, randomised 
controlled trial. The lancet HIV. 2020;7(5):e332-e9.  
10. 
Chinula L, Brummel SS, Ziemba L, Stranix-Chibanda L, Coletti A, Krotje C, et al. Safety and 
efficacy of DTG vs EFV and TDF vs TAF in pregnancy: IMPAACT 2010 trial.  Conference on Retroviruses 
and Opportunistic Infections; Boston, Mass. 2020. 
11. 
Zash R, Makhema J, Shapiro RL. Neural-Tube Defects with Dolutegravir Treatment from the Time 
of Conception. The New England journal of medicine. 2018;379(10):979-81.  
12. 
Zash R, Holmes L, Diseko M, Jacobson DL, Brummel S, Mayondi G, et al. Neural-Tube Defects 
and Antiretroviral Treatment Regimens in Botswana. The New England journal of medicine. 
2019;381(9):827-40.  
13. 
Raesima MM, Ogbuabo CM, Thomas V, Forhan SE, Gokatweng G, Dintwa E, et al. Dolutegravir 
Use at Conception - Additional Surveillance Data from Botswana. The New England journal of medicine. 
2019;381(9):885-7.  
14. 
Zash R, Holmes L, Diseko M, Jacobson D, Mayondi G, Isaacson A, et al. Update on neural tube 
defects with antiretroviral exposure in the Tsepamo study, Botswana.  23rd International AIDS Conference 
Virtual, 2020. 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 29 of 54 
15. 
Antiretroviral Pregnancy Registry Steering Committee. Antiretroviral Pregnancy Registry Interim 
Report for 1 January 1989 through 31 January 2020 Wilmington, NC: Registry Coordinating Center; 2020. 
Available from: www.APRegistry.com]. 
16. 
Amarin ZO, Obeidat AZ. Effect of folic acid fortification on the incidence of neural tube defects. 
Paediatric and perinatal epidemiology. 2010;24(4):349-51.  
17. 
De Wals P, Tairou F, Van Allen MI, Uh SH, Lowry RB, Sibbald B, et al. Reduction in neural-tube 
defects after folic acid fortification in Canada. The New England journal of medicine. 2007;357(2):135-42.  
18. 
Pereira GFM, Kim A, Jalil EM, Fernandes Fonseca F, Shepherd BE, Veloso VG, et al. Dolutegravir 
and pregnancy outcomes in women on antiretroviral therapy in Brazil: a retrospective national cohort study. 
The lancet HIV. 2021;8(1):e33-e41.  
19. 
Abrams E, Myer L. Lessons from dolutegravir and neural tube defects. The lancet HIV. 
2021;8(1):e3-e4.  
20. 
Santos LM, Lecca RC, Cortez-Escalante JJ, Sanchez MN, Rodrigues HG. Prevention of neural tube 
defects by the fortification of flour with folic acid: a population-based retrospective study in Brazil. Bulletin 
of the World Health Organization. 2016;94(1):22-9.  
21. 
Avagliano L, Massa V, George TM, Qureshy S, Bulfamante GP, Finnell RH. Overview on neural 
tube defects: From development to physical characteristics. Birth defects research. 2019;111(19):1455-67.  
22. 
Crider KS, Devine O, Hao L, Dowling NF, Li S, Molloy AM, et al. Population red blood cell folate 
concentrations for prevention of neural tube defects: Bayesian model. BMJ (Clinical research ed). 
2014;349:g4554.  
23. 
Dugdale CM, Ciaranello AL, Bekker LG, Stern ME, Myer L, Wood R, et al. Risks and Benefits of 
Dolutegravir- and Efavirenz-Based Strategies for South African Women With HIV of Child-Bearing 
Potential: A Modeling Study. Annals of internal medicine. 2019;170(9):614-25.  
24. 
Estill J, Bertisch B. More evidence for dolutegravir as first-line ART for all. The lancet HIV. 
2020;7(3):e154-e5.  
25. 
World Health Organization. Update of recommendations on first- and second-line antiretroviral 
regimens. 
Geneva, 
Switzerland: 
World 
Health 
Organization; 
2019. 
Available 
from: 
https://apps.who.int/iris/bitstream/handle/10665/325892/WHO-CDS-HIV-19.15-eng.pdf?ua=1]. 
26. 
Eswatini Ministry of Health. Swaziland Health Information Management System 2. 2017. 
27. 
Eswatini Ministry of Health. Annual HIV Programs Report. 2018. 
28. 
Central Statistical Office and UNICEF. Multiple Indicator Cluster Survey (MICS) Final Report 
2014. Mbabane, Eswatini: Central Statistical Office and UNICEF; 2016. Available from: https://mics-
surveys-
prod.s3.amazonaws.com/MICS5/Eastern%20and%20Southern%20Africa/Eswatini/2014/Final/Swaziland
%202014%20MICS%20Final%20Report_English.pdf]. 
29. 
EGPAF. Programmatic data. 2020. 
30. 
Watts DH. Teratogenicity risk of antiretroviral therapy in pregnancy. Current HIV/AIDS reports. 
2007;4(3):135-40.  
 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 30 of 54 
Participant PTID:  
Eswatini Birth Outcomes Study 
 
 
Assessment of Birth Outcomes in Eswatini after Transition to Dolutegravir-Based Treatment 
 
Chart Abstraction Form for live births, stillbirths and miscarriages 
 
MATERNITY NAME 
MBABANE GOVERNMENT HOSPITAL 
 1 
RALEIGH FITKIN MEMORIAL HOSPITAL 
 2 
GOOD SHEPHERD HOSPITAL 
 3 
HLATIKHULU HOSPITAL 
 4 
MANKAYANE GOVERNMENT HOSPITAL 
 5 
DATA COLLECTOR’S NAME/ID: 
 
NAME:  _________________________________ 
 
 ID: ___________ 
DAY / MONTH / YEAR OF ABSTRACTION: 
 
___ ___ /___ ___ /  2   0   ___ ___ 
DATA COLLECTION POINT:  
MATERNITY 
  
OPD/GYNECOLOGY 
 
 
 
# 
Variables 
Response Options  
Directions 
Pregnancy and HIV history, Source: registers 
1. 
Date of admission 
(dd/mm/yyyy) 
 
__ __ / __ __ / __ __ __ __ 
 
 
2. 
Mother’s age 
________________ years 
   1 
 Unknown/not documented 
 87 
 
3. 
Gravidity 
___ # of pregnancies  
   1 
Unknown/ not documented 
 87 
 
4. 
Parity 
___ # of births  
   1 
Unknown/not documented 
 87 
 
5. 
Last menstrual period (LMP) 
 
__________________________ 
 
 
6. 
Gestational age at L&D booking 
 
Not applicable for miscarriages/ 
medical induced abortions 
______ weeks, ______ days 
   1 
Don’t know/not documented 
 87 
N/A 
 99 
 
 
7. 
HIV status at L&D/admission 
 
 
HIV positive 
   1 
HIV negative 
   2 
Never tested for HIV 
   4 
Unknown/not documented 
 87 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 31 of 54 
8. 
Tested for HIV at L&D (indicate 
yes if there is a date of HIV test 
entered) 
 
Not applicable for miscarriages/ 
medically induced abortions 
 Yes 
   1  
No 
   2  
 Unknown/not documented 
 87  
 N/A 
 99 
 
SKIP TO Q 
SKIP TO Q 
SKIP TO Q 
9. 
HIV test result at L&D  
 
Not applicable for miscarriages 
HIV positive 
 10 
HIV negative 
 20 
Inconclusive 
 30 
N/A 
 99 
 
10. Date of most recent HIV test prior 
to L&D 
(dd/mm/yyyy) 
 
Not applicable for miscarriages 
__ __ / __ __ / __ __ __ __ 
   1  
Unknown/not documented 
 87 
N/A 
 99 
 
Birth outcomes, Source: registers 
11. Birth outcome  
 
Live birth  
 10 
Stillbirth  
 20 
Miscarriage 
 30 
Medically induced abortion 
 40 
 
 
If miscarriage or 
abortion, go to Q22 
12. Single or multiple birth 
 
Not applicable for miscarriages/ 
medically induced abortions 
Single  
 10 
Twin  
 20 
Triplet  
 30 
N/A 
 99 
For options 2 and 3, 
form will return to Q12 
to ask for each 
subsequent child while 
retaining the same 
maternal information 
13. Gestational age at delivery  
______ weeks, ______ days 
 10 
Unknown/not documented 
 87 
 
14. Mode of delivery 
Normal vaginal delivery (NVD) 
 10 
Assisted vaginal delivery (AVD) 
 20 
Elective C-section (ELCS) 
 30 
Emergency C-section (EMCS) 
 40 
 
15. Date of delivery 
(dd/mm/yyyy) 
 
__ __ / __ __ / __ __ __ __ 
 1  
 
 
16. If stillbirth 
FSB (fresh) 
   1 
MSB (macerated) 
   2  
N/A – live birth 
 99 
 
17. Sex of baby 
Male  
 10 
Female  
 20 
Ambiguous  
 30 
 
18. Weight at birth 
___ grams  
   1 
Unknown/not documented 
 87 
 
19. Neonatal death before discharge  
Yes 
 10 
No 
 20 
 
SKIP TO Q 
20. Causes of infant death 
___________________________  
   1 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 32 of 54 
Unknown/not documented 
 87 
21. Infant age at death 
______________________ hours 
   1 
Unknown/not documented 
 87 
For all responses go to 
Q25 
22. If miscarriage or abortion, date of 
miscarriage/abortion 
(dd/mm/yyyy) 
Same as date of admission 
    1  
Different from admission date:  
__ __ / __ __ / __ __ __ __ 
     2 
Unknown/not documented 
   87 
 
23. Gestational age at time of 
miscarriage/abortion 
___ # of births  
   1 
Unknown/not documented 
 87 
 
24. Pregnancy condition 
 
__________________________ 
 
 
25. Was a birth defect identified? 
Yes 
   1  
No 
   2  
 Unknown/not documented 
 87  
 N/A 
 99 
 
26. Comments: Please use this 
section to indicate any additional 
relevant information found in 
source documents, including 
maternal complications (e.g., 
preeclampsia, diabetes, etc.) or 
child’s condition (e.g., birth 
defect, jaundice, prematurity). 
 
Applies to all infants/birth 
outcomes. 
 
__________________________________________________________
__________________________________________________________
__________________________________________________________
__________________________________________________________
__________________________________________________________
__________________________________________________________ 
Antiretroviral Therapy (ART), Sources: registers, electronic database  
27. ART regimen 
Yes 
 1 
No 
 2 
 
SKIP to Q 
28. First two drugs (‘NRTI backbone’) 
listed in the CURRENT ART 
regimen: 
ABC/3TC 
   10 
AZT/3TC 
   20 
   TDF/3TC 
   30 
Other, specify:  ________________ 
   88   
Unknown/not documented 
   87 
 
 
 
 
 
 
29. Third drug listed in the CURRENT 
ART regimen:  
NVP 
   1 
EFV 
   2 
ATV/r 
   3 
LPV/r 
   4 
DTG 
   5 
DRV 
   6 
Other, specify: _________________ 
 88 
Unknown/not documented 
 87 
 
30. Date started CURRENT ART 
regimen 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
(dd/mm/yyyy) 
Unknown/not documented 
 87 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 33 of 54 
31. If unknown, when was current 
regimen started?  
During pregnancy 
   1 
Before pregnancy 
   2 
Unknown/not documented 
 87 
 
32. Changes in ART regimen in the 
past 12 months (from date of 
admission) 
Yes 
 1 
No 
 2 
 
SKIP to Q 
33. Number of changes in ART 
regimen in past 12 months 
One 
 1 
Two 
 2 
Three 
 3 
 
FIRST change in ART regimen in the last 12 months from date of admission. Must be different from CURRENT 
regimen. Source: electronic database 
34. First two drugs (‘NRTI backbone’) 
listed in the CURRENT ART 
regimen: 
ABC/3TC 
   1 
AZT/3TC 
   2 
TDF/3TC 
   3 
Other, specify:  ________________ 
 88 
Unknown/not documented 
 87 
 
 
 
 
35. Third drug listed in the CURRENT 
ART regimen:  
NVP 
   1 
EFV 
   2 
ATV/r 
   3 
LPV/r 
   4 
DTG 
   5 
DRV 
   6 
Other, specify: _________________ 
 88 
Unknown/not documented 
 87 
 
36. ART regimen start date 
(dd/mm/yyyy) 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
37. ART regimen stop date 
(dd/mm/yyyy) 
 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
38. Reason for stopping 
Side effects 
 10  
Treatment failure 
 20  
Drug interactions (e.g., TB treatment) 
 30  
Pregnancy 
 40  
Change in treatment protocol 
 50  
Other, specify: ____________ 
 88 
Unknown/not documented 
 87 
 
SECOND change in ART Regimen in the last 12 months from date of admission. Must be different from 
CURRENT regimen. Source: electronic database 
39. First two drugs (‘NRTI backbone’) 
listed in the CURRENT ART 
regimen: 
ABC/3TC 
   1 
AZT/3TC 
   2 
TDF/3TC 
   3 
Other, specify:  ________________ 
 88 
Unknown/not documented 
 87 
 
 
 
 
 
 
40. Third drug listed in the CURRENT 
ART regimen:  
NVP 
  1 
EFV 
  2 
ATV/r 
  3 
LPV/r 
  4 
DTG 
  5 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 34 of 54 
DRV 
   6 
Other, specify: _________________ 
 88 
Unknown/not documented 
 87 
41. ART regimen start date 
(dd/mm/yyyy) 
 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
42. ART regimen stop date 
(dd/mm/yyyy) 
 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
43. Reason for stopping 
Side effects 
 10  
Treatment failure 
 20  
Drug interactions (e.g., TB treatment) 
 30  
Pregnancy 
 40  
Change in treatment protocol 
 50  
Other, specify: ____________ 
 88 
Unknown/not documented 
 87 
 
THIRD change in ART Regimen in the last 12 months from date of admission. Must be different from CURRENT 
regimen. Source: electronic database 
44. First two drugs (‘NRTI backbone’) 
listed in the CURRENT ART 
regimen: 
ABC/3TC 
   1 
AZT/3TC 
   2 
TDF/3TC 
   3 
Other, specify:  ________________ 
 88 
Unknown/not documented 
 87 
 
45. Third drug listed in the CURRENT 
ART regimen:  
NVP 
   1 
EFV 
   2 
ATV/r 
   3 
LPV/r 
   4 
DTG 
   5 
DRV` 
   6 
Other, specify: _________________ 
 88 
Unknown/not documented  
 87 
 
46. ART regimen start date 
(dd/mm/yyyy) 
 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
47. ART regimen stop date 
(dd/mm/yyyy) 
 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
48. Reason for stopping 
Side effects 
 10  
Treatment failure 
 20  
Drug interactions (e.g., TB treatment) 
 30  
Pregnancy 
 40  
Change in treatment protocol 
 50  
Other, specify: ____________ 
 88 
Unknown/not documented 
 87 
 
Pre-exposure Prophylaxis (PrEP), Source: electronic database 
49. Receipt of PrEP in pregnancy 
(indicated from log) 
Yes 
   1 
No 
   2 
     Unknown/not documented 
 87 
 
END 
50. PrEP regimen 
TDF + 3TC  
   1 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 35 of 54 
Cabotegravir (injection) 
   2 
Dapivirine 
   3 
     Unknown/not documented 
 87 
51. Date started PrEP 
(dd/mm/yyyy) 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
52. Date stopped PrEP 
(dd/mm/yyyy) 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
53. Select yes if abortion 
Yes 
 10  
No 
 20 
 
Select ‘Yes’ if it’s an 
abortion and the 
woman is available for 
an interview. If yes, 
conduct entire 
interview.  
 
Select “No” if it is an 
abortion and women is 
not available for an 
interview. If “No”, skip 
to PART V: BIRTH 
DEFECT QUESTIONS on 
the women interview 
form. 
 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 36 of 54 
Eswatini Birth Outcomes Study  
Participant Number: 
 
 
 
 
Assessment of Birth Outcomes in Eswatini following Transition to Dolutegravir-based Treatment 
 
Women Interview Form for Suspected Birth Defects 
 
MATERNITY NAME 
MBABANE GOVERNMENT HOSPITAL 
 1 
RALEIGH FITKIN MEMORIAL HOSPITAL 
 2 
GOOD SHEPHERD HOSPITAL 
 3 
HLATIKHULU HOSPITAL 
 4 
MANKAYANE GOVERNMENT HOSPITAL 
 5 
INTERVIEWER’S NAME/ID: 
 
NAME:  _________________________________ 
 
 ID: ___________ 
DAY / MONTH / YEAR OF INTERVIEW: 
 
___ ___ /___ ___ /  2   0   ___ ___ 
INDICATE IF; 
LIVE/STILLBIRTH, START FROM PART 1 
MEDICAL ABORTION, WOMEN AVAILABLE, START FROM PART 1 
MEDICAL ABORTION, WOMEN NOT AVAILABLE GO TO PART V: 
 
 
PART I.  MATERNAL CHARACTERISTICS AND HEALTH HISTORY  
 
Instructions: I will start with questions about your pregnancy history. I will ask you questions but also verify in your records 
as we go along. 
1.1 
 
Nationality  
Eswatini  
 10 
Other, specify: ____________ 
 88 
 
 
1.2 
Date of last menstrual period 
(dd/mm/yyyy) 
From records: ___/___/_____ 
 11  
Estimated: ___/___/_____ 
 21 
 
1.3 
Gestational age at booking 
______ weeks, ______ days
   1 
Don’t know/not documented 
 87 
 
1.4 
Mother’s weight at first ANC  
_____ Kgs  
 10 
Don’t know 
 87 
 
1.5 
Mother’s most recent weight  
At last ANC _____ Kgs  
 11 
At delivery _____ Kgs  
 21 
Don’t know at both ANC and delivery  
 87 
 
1.6 
Mother’s height at ANC 
___ Cm  
 10 
Don’t know 
 87 
 
1.7 
Gravida 
___ # of pregnancies  
 10 
Don’t know 
 87 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 37 of 54 
1.8 
Parity 
___ # of births  
 10 
Don’t know 
 87 
 
1.9 
Have you had any previous preterm 
deliveries (about < 37 weeks gestation)? 
Yes 
 10 
No 
 20 
Don’t know 
 87 
N/A 
 99 
 
1.10 
Have you had any previous stillborn 
children? 
Yes 
 10 
No 
 20 
Don’t know 
 87 
N/A 
 99 
 
1.11 
How many birth defects have you had in 
your previous pregnancies? 
None 
 10 
Number: ____ 
 20 
Don’t know 
 87 
N/A 
 99 
 
1.12 
If yes, please specify birth defect(s) or 
describe (if name unknown) 
_________________________________ 
 
 
HIV-RELATED HISTORY  
1.13 
When was your most recent HIV test 
done? 
Before pregnancy 
 10 
During pregnancy, before delivery 
 20 
At labor and delivery 
 30 
After delivery, before hospital discharge 
 40 
Don’t know 
 87 
 
1.14 
What is your HIV status? 
 
 
HIV positive 
 10 
HIV negative 
 20 
Don’t know 
 87 
Never tested for HIV 
 40 
 
SKIP TO Q 
SKIP TO Q 
SKIP TO Q 
1.15 
When was your first positive HIV test? 
 ___/___/_____ (dd/mm/yyyy) 
 10  
Don’t know 
 87 
 
1.16 
When was your most recent HIV negative 
test? 
___/___/_____ (dd/mm/yyyy) 
 10  
Don’t know 
 87 
 
1.17 
When did you initiate ART?  
___/___/_____ (dd/mm/yyyy) 
 10  
Don’t know 
 87 
N/A – never on ART  
 99 
 
 
 
PART II.  HIV-RELATED DRUG USE 
 
Instructions (to say to participant): For this and subsequent questions, we are interested in learning more about what 
health conditions you have and what medicines you took before and after pregnancy.  I will start with asking about HIV 
medicines, for treatment or prophylaxis. Then, I will ask about any medical conditions and other medicines, selected 
behaviors and any pregnancy complications. Finally, I will talk to you about the baby and birth and information on the 
suspected birth defect. 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 38 of 54 
For interviewers, please note: Before pregnancy refers to the time from the month before pregnancy based on LMP or best 
estimate. This section should be skipped for HIV negative women.  
2.1 
 
Is there documentation of ANY ARVs 
initiated or continued during this 
pregnancy? 
Yes 
 10 
No 
 20 
 
SKIP TO Q 
2.2 
Is there documentation of ARVs at the 
time of conception? 
Yes, ART 
 11  
Yes, PrEP 
 21  
No 
 31 
Don’t know/not documented 
 87 
 
If ‘2’ Skip to 
Q2.32 
2.3 
Initial regimen prescribed or continued 
during pregnancy 
TDF+3TC 
 10  
TDF+3TC+EFV 
 20  
TDF+3TC+DTG 
 30  
AZT+3TC+DTG 
 40  
AZT+3TC+EFV 
 50  
Other, specify: ____________ 
 88  
 
2.4 
Start date of initial regimen: 
dd/mm/yyyy 
  __ __ / __ __ / __ __ __ __ 
 
2.5 
Indicate if any part of the date is 
estimated (can select more than one if a 
‘Yes’ response is selected) 
No 
 10 
Yes, day 
 20 
 Yes, month 
 30 
Yes, year 
 40 
 
2.6 
Is there documentation of change of 
ARVs to a new regimen during 
pregnancy? 
Yes 
 10 
No 
 20 
 
SKIP TO Q 
2.7 
How many times were ARVs changed 
during pregnancy? 
One 
 10  
Two 
 20  
 Three 
 30  
 
2.8 
Date of first change of ARV regimen: 
dd/mm/yyyy 
__ __ / __ __ / __ __ __ __ 
 
2.9 
Indicate if any part of the date is 
estimated (can select more than one if a 
‘Yes’ response is selected). 
No 
 10 
Yes, day 
 20 
 Yes, month 
 30 
Yes, year 
 40 
 
2.10 
Reason for first change of ARV regimen? 
 
Side effects 
 10  
Treatment failure 
 20  
Drug interactions (e.g., TB treatment) 
 30  
Pregnancy 
 40  
Change in treatment protocol 
 50  
Other, specify: ____________ 
 88 
Don’t know/not documented 
 87 
 
2.11 
Second ARV regimen in pregnancy? 
TDF+3TC+EFV 
 10  
TDF+3TC+DTG 
 20  
AZT+3TC+DTG 
 30  
AZT+3TC+EFV 
 40  
Other, specify: ____________ 
 88 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 39 of 54 
2.12 
Date of second change of ARV regimen: 
dd/mm/yyyy 
__ __ / __ __ / __ __ __ __ 
 
2.13 
Indicate if any part of the date is 
estimated (can select more than one if a 
‘Yes’ response is selected) 
No 
 10 
Yes, day 
 20 
 Yes, month 
 30 
Yes, year 
 40 
 
2.14 
Reason for second change of ARV 
regimen? 
Side effects 
 10  
Treatment failure 
 20  
Drug interactions (e.g., TB treatment) 
 30  
Pregnancy 
 40  
Change in treatment protocol 
 50  
Other, specify: ____________ 
 88 
Don’t know/not documented 
 87 
 
2.15 
Third ARV regimen in pregnancy? 
TDF+3TC+EFV 
 10 
TDF+3TC+DTG 
 20  
AZT+3TC+DTG 
 30 
AZT+3TC+EFV 
 40  
Other, specify: ____________ 
 88 
 
2.16 
Date of third change of ARV regimen: 
dd/mm/yyyy 
__ __ / __ __ / __ __ __ __ 
 
2.17 
Indicate if any part of the date is 
estimated (can select more than one if a 
‘Yes’ response is selected) 
No 
 10 
Yes, day 
 20 
 Yes, month 
 30 
Yes, year 
 40 
 
2.18 
Reason for third change of ARV regimen? 
Side effects 
 10  
Treatment failure 
 20  
Drug interactions (e.g., TB treatment) 
 30  
Pregnancy 
 40  
Change in treatment protocol 
 50  
Other, specify: ____________ 
 88 
Don’t know/not documented 
 87 
 
2.19 
Fourth ARV regimen in pregnancy? 
TDF+3TC+EFV 
 10 
TDF+3TC+DTG 
 20  
AZT+3TC+DTG 
 30 
AZT+3TC+EFV 
 40  
Other, specify: ____________ 
 88 
 
2.20 
Date of fourth change of ARV regimen: 
dd/mm/yyyy 
__ __ / __ __ / __ __ __ __ 
 
2.21 
Indicate if any part of the date is 
estimated (can select more than one if a 
‘Yes’ response is selected) 
No 
 10 
Yes, day 
 20 
 Yes, month 
 30 
Yes, year 
 40 
 
2.22 
During the time of your last menstrual 
period, did you miss any days taking your 
ARVS? 
Yes 
 10 
No 
 20 
Don’t know 
 87 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 40 of 54 
2.23 
Is there documentation of termination or 
defaulting of ARVs during pregnancy? 
(stopping ARVs and not starting onto a 
new regimen prior to delivery) 
 
Yes responses include women not on 
ARVs at time of delivery but previously on 
ARVs in pregnancy 
Yes  
 10 
No 
 20 
 
SKIP TO Q 
2.24 
Date of ARV termination/defaulting date: 
dd/mm/yyyy 
 __ __ / __ __ / __ __ __ __ 
 
2.25 
Indicate if any part of the date is 
estimated (can select more than one if a 
‘Yes’ response is selected). 
No 
 10 
Yes, day 
 20 
 Yes, month 
 30 
Yes, year 
 40 
 
2.26 
Comments (please provide any ARV 
related clarifications or explanations for 
a disruption of ARVs >1 week). 
__________________________________
__________________________________
__________________________________
__________________________________
__________________________________ 
__________________________________
__________________________________
__________________________________ 
__________________________________
__________________________________ 
 
2.27 
Did you take any other medicine 
(including traditional medicine) or 
therapy? 
Yes 
 10 
No 
 20 
 
SKIP TO Q 
2.28 
If yes, first medicine/therapy: 
____________________ 
 
2.29 
First medicine/therapy: When did you 
take it? (multiple responses permitted in 
some cases)  
Up to a month before pregnancy 
 10 
In 1st trimester 
 20 
In 2nd trimester 
 30  
In 3rd trimester 
 40  
Don’t know if before or during pregnancy 
 50   
During pregnancy, but don’t know trimester  
 60 
 
2.30 
If yes, second medicine/therapy: 
_____________________ 
No second therapy 
 99 
 
SKIP TO Q 
2.31 
Second medicine/therapy: When did you 
take it? (multiple responses permitted in 
some cases)  
Up to a month before pregnancy 
 10 
In 1st trimester 
 20 
In 2nd trimester 
 30  
In 3rd trimester 
 40  
Don’t know if before or during pregnancy 
 50   
During pregnancy, but don’t know trimester  
 60 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 41 of 54 
2.32 
If on PrEP, PrEP regimen 
TDF+3TC 
   1 
Cabotegravir (injection) 
   2 
Dapivirine 
   3 
     Unknown/not documented 
 87 
 
2.34 
Date started PrEP 
(dd/mm/yyyy) 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
2.35 
Date stopped PrEP 
(dd/mm/yyyy) 
Known 
   1 
Date: __ __ / __ __ / __ __ __ __ 
     Unknown/not documented 
 87 
 
 
 
 
PART III. RISK FACTORS 
 
MICRONUTRIENTS 
 
Micronutrient Use 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.1 
FOLIC ACID 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.2 
MULTIVITAMIN   
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.3 
FERROUS SULFATE 
(IRON) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
 
DIABETES 
3.4 
Do you currently have diabetes? 
Yes 
 10 
No  
 20 
Don’t know  
 87 
 
  
3.5 
Did you have diabetes before this pregnancy? 
Yes 
 10 
No  
 20 
Don’t know  
 87 
 
3.6 
If yes to 3.4, what medicines for your diabetes 
did you take very early in this pregnancy 
between your first and third month of 
pregnancy? (Response should exclude 
traditional medicines) 
None 
 11 
Injections  
 21 
Tablets  
 31 
 
3.7 
If yes to 3.4, were you able to keep your sugar 
in control during this pregnancy?  
Yes 
 10 
No  
 20 
Don’t know  
 87 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 42 of 54 
 
CONVULSIONS (EPILEPSY) 
3.8 
Have you ever had convulsions? 
Yes 
 10 
No  
 20 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 
3.9 
Did you have convulsions just before you got pregnant? 
Yes 
 10 
No  
 20 
Don’t know  
 87 
 
3.10 
 
Did you have any convulsions during this pregnancy? 
Yes 
 10 
No  
 20 
Don’t know  
 87 
 
 
 
 
Medicines used for 
convulsions 
(Epilepsy) 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.11 
VALPROIC ACID 
(DEPAKENE) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.12 
PHENYTOIN (DILANTIN)   
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.13 
FELBAMATE (FELBATOL) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.14 
CARBAMAZEPINE 
(TEGRETOL) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.15 
PHENOBARBITAL 
(LUMINAL) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.16 
OTHER MEDICINE, 
SPECIFY: 
________________ 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
 
MALARIA 
3.17 
Did you have malaria during this pregnancy? 
Yes 
 10 
No  
 20 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 
3.18 
Did you take any medicines or remedies for 
malaria during this pregnancy? 
Yes 
 10 
No  
 20 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 43 of 54 
 
Medicines used for 
Malaria 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.19 
COARTEM  
(ARTEMETHER/ 
LUMAFANTRINE) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.20 
DUOCOTEXIN 
(DIHYDROARTEMISININ/ 
PIPERAQUINE) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.21 
FANSIDAR 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.22 
QUININE 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.23 
UNKNOWN MEDICINE 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.24 
OTHER MEDICINE, 
SPECIFY: 
_________________ 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
 
TUBERCULOSIS 
3.25 
Did you have tuberculosis during this 
pregnancy? 
Yes 
 11 
No  
 21 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 
3.26 
Did you take any medications or remedies for 
tuberculosis during this pregnancy? 
Yes 
 11 
No  
 21 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 
 
Medicines used for 
TB 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.27 
INH 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.28 
RIFAMPIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 44 of 54 
 
Medicines used for 
TB 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.29 
PYRAZINAMIDE (PZA) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.30 
ETHAMBUTOL 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.31 
STREPTOMYCIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.32 
UNKNOWN MEDICINE 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.33 
OTHER MEDICINE, 
SPECIFY: 
_________________ 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
 
FEVER 
3.34 
Did you have any fevers, including those due to 
malaria, TB, flu with difficulty breathing/bad 
cough, pneumonia, or other illness? 
Yes 
 11 
No  
 21 
Don’t know  
 87 
 
 
 
3.35 
Did you have fevers around the time you got 
pregnant?   
Yes 
 11 
No  
 21 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 
3.36 
Did you take any medicines or remedies for the 
fever? 
 
Yes 
 11 
No  
 21 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 
 
Medicines used for 
Fever 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.37 
ASPIRIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.38 
PARACETAMOL 
(PANADOL) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.39 
IBUPROFEN (BRUFEN) 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 45 of 54 
 
Medicines used for 
Fever 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.40 
DICLOFENAC 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.41 
OTHER MEDICINE, 
SPECIFY: 
_________________ 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
 
ANTIBIOTIC USE 
3.42 
Did you take antibiotics for anything other than 
fever during this pregnancy? 
Yes 
 11 
No  
 21 
Don’t know  
 87 
 
SKIP TO Q 
SKIP TO Q 
3.43 
If yes, for what condition? 
Specify: ______________________
 11 
Don’t know  
 87 
 
 
Antibiotics 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.44 
CIPROFLOIXACIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.45 
CEFTRIAXONE 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.46 
ERYTHROMYCIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.47 
AMOXICILLIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.48 
DOXYCYCLINE 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.49 
PENICILLIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.50 
CLOXACILLIN 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 46 of 54 
 
Antibiotics 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.51 
FLUCONAZOLE 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.52 
 COTRIMOXAZOLE 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.53 
OTHER, SPECIFY: 
_________________ 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but don’t 
know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
 
ANY OTHER DRUG/THERAPY USE 
 
Please use the space below to document any other drugs or therapies that were taken before pregnancy (= time 
from the month before pregnancy based on LMP or best estimate) or during pregnancy. 
 
Other 
No 
Yes, but unknown timing 
Yes, Before 
pregnancy 
Yes, 1st 
trimester 
Yes, 2nd 
trimester 
Yes, 3rd 
trimester  
3.54 
_________________ 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
3.55 
_________________ 
☐ 
Don’t know if before or 
during pregnancy 
 1 
During pregnancy, but 
don’t know trimester 
 2 
☐ 
☐ 
☐ 
☐ 
 
TRADITIONAL MEDICINE 
3.56 
Did you take any traditional medicine right 
before pregnancy or at any time during 
pregnancy? 
Yes 
 10 
No 
 20 
Don’t know 
 87 
 
SKIP TO Q 
SKIP TO Q 
3.57 
If yes, what condition did you take it for?  
 
Select all that apply. The next two questions 
will be asked for each condition indicated 
here.  
Diabetes 
 10 
Convulsions (Epilepsy) 
 20 
Malaria 
 30 
TB 
 40 
Fever 
 50 
Other, specify: _________________ 
 88 
Don’t know 
 87 
 
 
3.58 
Indicate the name or main ingredient of the 
medicine. 
Specify: _____________________________ 
 100 
Don’t know 
 870 
 
3.59 
When did you take it?  
 
 
During first trimester 
 10 
During second trimester 
 20 
During third trimester 
 30 
Before pregnancy 
 40 
    During pregnancy, but don’t know trimester 
 50 
May have 
multiple 
responses 
to 1-5; if 
select 6, 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 47 of 54 
Don’t know if before or during pregnancy 
 60 
 
should be 
only 
response  
 
 
BEHAVIORAL RISK FACTORS  
3.60 
How often did you consume alcohol during 
pregnancy?  
Never 
 10 
Occasionally 
 20 
Sometimes 
 30 
Frequently 
 40 
 
3.61 
How often did you smoke cigarettes or use 
other tobacco products during pregnancy?  
Never 
 10 
Occasionally 
 20 
Sometimes 
 30 
Frequently 
 40 
 
 
PREGNANCY COMPLICATIONS AND COVID-19 
3.62 
Maternal diagnoses/complications during 
pregnancy (select all that apply) 
 
 
None 
 10 
Pre-eclampsia 
 20 
Hypertension 
 30 
Diabetes mellitus 
 40 
Excess weight gain, obesity 
 50 
Syphilis 
 60 
High blood pressure 
 70 
Other, specify: _________________ 
 88 
 
3.63 
Hemoglobin at ANC  
 Hgb at first ANC: _______ 
 10  
Hgb at other ANC visit (if 1st  not available): 
_______ 
 20 
Don’t know  
 87 
 
3.64 
Were you diagnosed with COVID-19 
during this pregnancy?  
 
Yes 
 10 
No 
 20 
 
 
SKIP TO Q 
3.65 
If yes, how was it diagnosed?  
COVID-19 test done 
 10 
Diagnosis based on symptoms only 
 20 
Other, specify: _________________ 
 88 
 
3.66 
If yes, what trimester were you 
diagnosed? 
First 
 10 
Second 
 20 
Third 
 30 
Don’t know 
 87 
 
 
 
 
PART IV. INFANT AND DELIVERY OUTCOMES 
 
 
Instructions: I will now ask you questions about the birth (and child if NOT still born). As before, I will ask you 
questions while also verifying in your records.  
4.1 
Gestational age at delivery (or at time of 
medical induced abortion) 
______ weeks, ______ days
   1 
Don’t know/not documented 
 87 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 48 of 54 
4.2 
Single or multiple birth 
Single  
 10 
Twin  
 20 
Triplet  
 30 
Qs should repeat 
for second and 
third births 
4.3 
Date of birth or abortion (dd/mm/yyyy) 
 
__ __ / __ __ / __ __ __ __ 
 
 
4.4 
Type of birth 
Liveborn 
 10 
Stillborn  
 20 
Medically Induced Abortion  
 30 
 
4.5 
Birth weight 
___ grams  
 10 
Don’t know 
 87 
 
4.6 
Child sex 
Male  
 10 
Female  
 20 
Ambiguous  
 30 
For abortion and 
stillbirth, Q should 
be answered, then 
skip to Q4.11 
4.7 
Length 
___ cm  
 10 
Don’t know 
 87 
 
4.8 
Head circumference  
___ cm  
 10 
Don’t know 
 87 
 
4.9 
Apgar score 
_______ 
 10 
Don’t know 
 87 
 
4.10 
Newborn complications  
Jaundice 
 10 
Hypoglycemia 
 20 
Resuscitation 
 30 
Other, specify: _____________ 
 88 
 
4.11 
Date birth defect identified 
(dd/mm/yyyy) 
 
__ __ / __ __ / __ __ __ __ 
 
 
 
PART V. BIRTH DEFECT QUESTIONS 
 
5.1 
One or more abnormalities identified:  
 
First infant born 
 10 
Second infant born 
 20 
Third infant born 
 30  
 
Skip pattern to go 
to the affected 
child(ren) 
5.2 
Is the abnormality an extra digit? 
Yes 
 10 
No 
 20 
 
SKIP TO Q 
5.3 
Where are the extra digits? (select all that 
apply) 
Left hand 
 10 
Right hand 
 20 
Left foot 
 30 
Right foot 
 40 
Don’t know 
 87 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 49 of 54 
5.4 
Where on the left hand is the extra digit 
located? 
Near the fifth digit (pinky finger) 
 10 
Near the thumb 
 20 
Other place, specify: ___________ 
 88 
 
5.5 
Does the extra digit appear as: 
A fullly formed extra digit (seems to look like 1 
a finger and have bone in it) 
 10 
Soft, without a bone (sometimes hanging by 1 
a stalk) 
 20 
 
5.6 
Where on the right hand is the extra digit 
located? 
Near the fifth digit (pinky finger) 
 10 
Near the thumb 
 20 
Other place, specify: ___________  
 88 
 
5.7 
Does the extra digit appear as: 
A fullly formed extra digit (seems to look like 1 
a finger and have bone in it) 
 10 
Soft, without a bone (sometimes hanging by 1 
a stalk) 
 20 
 
5.8 
Where on the left foot is the extra digit 
located? 
Near the fifth digit (pinky toe) 
 10 
Near the big toe 
 20 
Other place, specify: ____________  
 88 
 
5.9 
Does the extra digit appear as: 
A fullly formed extra digit (seems to look like1   
a finger and have bone in it) 
 10 
Soft, without a bone (sometimes hanging by a1 
stalk) 
 20 
 
5.10 
Where on the right foot is the extra digit 
located? 
Near the fifth digit (pinky toe) 
 10 
Near the big toe 
 20 
Other place, specify: ____________ 
 88 
 
5.11 
Does the extra digit appear as: 
A fullly formed extra digit (seems to look like 1 
a finger and have bone in it) 
 10 
Soft, without a bone (sometimes hanging by 1 
a stalk) 
 20 
 
5.12 
Please provide any other details on this 
abnormality that are not captured above.  
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.13 
Is the head and neck exam NORMAL? 
(includes skull, fontanelles, eyes, ears, 
nose, jaw) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 50 of 54 
5.14 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.15 
Is there evidence of hydrocephalus 
(swollen head)? 
Yes 
 10 
No 
 20 
Don’t know 
 87 
 
5.16 
Is there a defect in the skull? 
Yes 
 10 
No 
 20 
Don’t know 
 87 
 
5.17 
If there is a defect in the skull, is it:  
Completely covered by skin 
 10 
Partly covered by skin 
 20 
Covered by a thin sac/membrane
 30 
Not covered by skin (brain tissue visible) 
 40 
Don’t know 
 87 
 
5.18 
Is the mouth, lip and palate exam 
NORMAL? (cleft lip or palate) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 
5.19 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.20 
Is the chest exam NORMAL? (shape and 
respiratory movements) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 
5.21 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.22 
Are the abdominal and anal exam 
NORMAL? (masses/anal 
closure defect) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 51 of 54 
5.23 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.24 
Are the arms and legs NORMAL? (length, 
shape, missing parts) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 
5.25 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.26 
Are the fingers and toes NORMAL? 
(including nails, number, dangling fused, 
shape or parts missing, abnormally large 
or small) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 
5.27 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.28 
Is the Spine exam NORMAL? (lumps or 
cysts or bulging in the back including the 
neck, thorax or lumbar area) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 
5.29 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If unknown, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 52 of 54 
5.30 
Where in the spine is the defect located? 
(Select all that apply) 
Cervical 
 10 
Thoracic 
 20 
Lumbosacral 
 30 
Don’t know 
 87 
 
5.31 
Is the spinal defect:  
Covered in hair and NOT bulging out 
 10 
Covered in hair and bulging out 
 20 
Bulging out and completely covered with skin1
 30  
Bulging out and partly covered with skin
 40  
Bulging out and covered with only a 1  
sac/thin membrane 
 50 
Open-can see the spine/spinal cord 
 60  
Don’t know 
 87 
 
5.32 
Are the hips and genitalia NORMAL? 
(including urethra, testes, penile shaft, 
vagina, labia) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 
5.33 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.34 
Is the skin exam NORMAL? (pale, blue, 
birth marks, or any large very red areas) 
Yes 
 10 
No 
 20 
Don’t know 
 87 
SKIP TO Q 
5.35 
If no, please describe abnormality: 
(please include as much detail as 
possible) 
 
If not known, explain why: 
 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 
5.36 
Describe any other abnormality, unusual 
finding or make any comment about 
abnormality 
Description: ____________________________ 
______________________________________
______________________________________
______________________________________ 
______________________________________
______________________________________
______________________________________
______________________________________
______________________________________ 
 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26AUG2022 
Page 53 of 54 
5.37 
Please indicate the preliminary diagnosis 
for this child. 
Central nervous system: 
Anencephaly 
  11 
Encephalocele 
  21  
Microcephaly 
  31 
Myelomeningocoele/Spina bifida, open0 
defect 
  41 
Myelomeningocoele/Spina bifida, closed0 
defect 
  51  
Hydrocephalus 
  61 
Craniorachischisis
  71  
Eye and ear malformations: 
Anophthalmia 
  81 
Microphthalmia 
  91 
Anotia 
 101  
Microtia 
 111 
Orofacial clefts: 0 
Cleft lip 
 120 
Cleft palate 
 130 
Cleft lip and palate 
 140  
Malformations of digestive system and 
genital organs 
Imperforate anus 
 150 
Hypospadias 
 160 
Musculoskeletal: 
Talipes equinovarus / Clubfoot 
 170 
Reduction defects of the limb(s) 
 180 
Exomphalos (omphalocele) 
 190 
Gastroschisis 
 200 
Polydactyly 
 210 
 
Other, specify: __________________ 
  88 
 
 
5.38 
Confirm that a doctor has reviewed and 
approved of the responses in section 5.  
Yes 
 10 
No 
 20 
If no, why? ___________________________ 
 
5.39 
Providers completing or providing input 
on this form (list all that apply) 
Doctor name: __________________ 
Midwife name: _________________ 
Other: ______________________________ 
 

Eswatini Birth Outcomes Study Protocol_Ver 3.0_26 AUG 2022 
Page 54 of 54