Conditions: Childhood Mortality
Keywords: childhood mortality neonates azithromycin mass drug administration
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 21832
Healthy volunteers: yes

Brief summary:
Although under-5 mortality rates are declining globally, neonatal mortality remains persistently high in many regions of sub-Saharan Africa. Mass azithromycin distribution to children aged 1-59 months has been shown to reduce childhood mortality in Niger, Tanzania, and Malawi. This study did not evaluate the effect of azithromycin administered during the neonatal period. Observational evidence from high income countries has suggested that macrolides, including erythromycin and azithromycin, may be associated with increased risk of development of infantile hypertrophic pyloric stenosis (IHPS). However, these studies are limited by confounding by indication, as infants only receive antibiotics when they are ill.

The investigators proposed an individually randomized trial of azithromycin versus placebo to establish the efficacy and safety of administration of a dose of azithromycin during the neonatal period. The long-term goal is generate evidence that can be used by neonatal and child survival programs related to the use of azithromycin in the youngest children who have the highest risk of mortality. The investigators hypothesize that a single dose of azithromycin administered in the neonatal period will lead to significantly reduced risk of mortality and that this dose will be safe.

Objectives

1. Establish the efficacy of a single dose of azithromycin administered during the neonatal period compared to placebo in infants 8 to 27 days of life for reduction in all-cause mortality.
2. Establish the safety of a single dose of azithromycin administered during the neonatal period.

This study will be conducted in several regions of Burkina Faso, including peri-urban areas of Ouagadougou and Nouna town, and rural areas that are within 4 hours' drive of a pediatric facility with capacity for performing pyloromyotomy

Detailed description:
Child mortality in West Africa is among the highest in the world. Although child health and mortality are improving worldwide, children in the Sahel and sub-Sahel regions of West Africa have the greatest risks of mortality. Burkina Faso's current under-5 mortality rate is estimated 110 per 1,000 live births. Similar to other countries in the region, the major causes of child mortality in Burkina Faso are malaria, respiratory tract infection, and diarrhea. Malnutrition acts as a major underlying contributor to mortality. Neonatal mortality remains persistently high, with approximately 1/5th of neonatal mortality due to pneumonia, meningitis, and sepsis. Interventions that address these underlying causes may be particularly efficacious for reducing mortality.

Younger children at are at a higher risk of mortality. Approximately 2/3rd of under-5 deaths occur during the first year of life. In general, the child mortality rate decreases as age increases. While some improvement has been observed, neonatal mortality is declining at a slower rate than post-neonatal childhood mortality. Many child health interventions are designed specifically for children over 6 months of age, such as vitamin A supplementation, seasonal malaria chemoprevention, and lipid-based nutritional supplementation. Identification of strategies that are safe and effective for the youngest children will be required to address persistently high rates of neonatal and infant mortality.

The MORDOR I study demonstrated a significant reduction in all-cause child mortality following biannual mass azithromycin distribution. Across three diverse geographic locations in sub-Saharan Africa (Malawi, Niger, and Tanzania), biannual mass azithromycin distribution over a two-year period led to a 14% decrease in all-cause child mortality. In Niger, 1 in 5-6 deaths were averted. These results are qualitatively similar to those of a previous study of mass azithromycin distribution for trachoma control in Ethiopia, which found reduced odds of all-cause mortality in children in communities receiving mass azithromycin compared to control communities.

In MORDOR I, the strongest effect of azithromycin was in the youngest cohort of children. Across all three countries, the strongest effect of azithromycin was consistently in children 1-5 months of age, with an approximately 25% reduction in all-cause mortality. However, MORDOR I was not optimized to target the youngest age groups. Although children as young as 1 month were eligible, biannual distributions might not reach some children until 7 months of age. On average, children were first treated at 4 months. Given that there may be a substantial benefit to treating children at younger ages, azithromycin strategies that are designed to target younger age groups may be even more beneficial for reducing child mortality.

Here, the investigators propose a randomized controlled trial designed to evaluate the efficacy of a dose of azithromycin administered during the neonatal period for prevention of mortality within in the first 6 months of life. The investigators propose to randomize births in several geographic regions of Burkina Faso to a single dose of azithromycin or placebo between day 8 and 27 of life. This study is designed to provide evidence of the efficacy of azithromycin treatment for the youngest children.

Arms:
arm 1: a single dose of Azithromycin will be administered to infants between their 8-27th days of life arm 2: a single dose of placebo will be administered to infants between their 8-27th days of life

Interventions: intervention 1: Azithromycin intervention 2: Placebo
Intervention descriptions: intervention 1: a single dose of Azithromycin will be administered to infants between their 8-27th days of life intervention 2: a single dose of Placebo will be administered to infants between their 8-27th days of life

Protocol document:
Prot_000.pdf: 
Version 1.3 - August 
2021 
1 
 Neonates and Azithromycin, an Innovation 
in the Treatment of Children in Burkina Faso 
NAITRE 
Manual of Operations and Procedures 
Centre de Recherche en Santé de Nouna 
The Francis I. Proctor Foundation, Global Health Sciences 
University of California, San Francisco 
 
Contents 
Abbreviations ................................................................................................................................... 4 


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2 
1.
Chapter 1: Overview ............................................................................................................... 5 
1.1 
Executive summary .......................................................................................................... 5 
1.2 
Objectives ......................................................................................................................... 5 
1.3 
Study Sites ........................................................................................................................ 5 
2 
Chapter 2: Context .................................................................................................................. 6 
3 
Chapter 3: Study Design ......................................................................................................... 7 
3.1 
Recruitment ...................................................................................................................... 7 
3.1.1 
Antenatal clinic recruitment .................................................................................... 8 
3.1.2 
Facility birth recruitment ........................................................................................ 8 
3.1.3 
Key informant-based recruitment ........................................................................... 8 
3.2 
Enrollment ........................................................................................................................ 8 
3.3 
Randomization ................................................................................................................. 8 
3.3.1 
Unit of randomization ............................................................................................. 8 
3.3.2 
Randomizing treatment ........................................................................................... 8 
4 
Chapter 4: Eligibility ............................................................................................................. 10 
4.1 
Eligible Communities ..................................................................................................... 10 
4.2 
Eligible Participants ....................................................................................................... 10 
5 
Chapter 5: Procedures ........................................................................................................... 12 
5.1 
Treatment ....................................................................................................................... 12 
5.2 
Field worker safety assessment ...................................................................................... 13 
5.3 
Vital status assessment ................................................................................................... 13 
5.4 
Anthropometry assessment ............................................................................................ 13 
5.5 
Passive Surveillance ....................................................................................................... 13 
5.6 
Long term follow up ....................................................................................................... 14 
6 
Chapter 6: Adverse Event Monitoring and Safety Assessment ............................................ 19 
6.1 
Background on Infantile Hypertrophic Pyloric Stenosis in Neonates ............................ 19 
6.2 
Screening for IHPS ......................................................................................................... 20 
7 
Chapter 7: Training ............................................................................................................... 22 
8 
Chapter 8: Study Medication ................................................................................................ 22 
8.1 
Study Medication Description (from Pfizer, Inc.) .......................................................... 22 
8.2 
Dosage Information ........................................................................................................ 22 
8.3 
Medication Procurement/Donation ................................................................................ 22 
8.4 
Medication Quality Control ........................................................................................... 22 
8.5 
Study Treatment ............................................................................................................. 23 


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8.6 
Serious Adverse Events .................................................................................................. 23 
8.7 
Adverse Events Data ...................................................................................................... 24 
9 
Chapter 9: Protection of Human Subjects ............................................................................. 25 
9.1 
Institutional Review Board Approval ............................................................................ 25 
9.2 
Informed Consent ........................................................................................................... 25 
10 
Chapter 10: Data and Safety Monitoring Committee Charter .......................................... 26 
10.1 Primary Responsibilities of the DSMC .......................................................................... 26 
10.2 DSMC Membership ....................................................................................................... 27 
10.3 Conflicts of Interest ........................................................................................................ 27 
10.4 Timing and Purpose of the DSMC Meetings ................................................................. 27 
10.5 Procedures to Ensure Confidentiality and Proper Communication ............................... 27 
10.6 Statistical Monitoring Guidelines ................................................................................... 29 
11 
Chapter 11: Data Collection, Management, and Security ................................................. 30 
11.1 Scope of Data ................................................................................................................. 30 
11.2 Data Storage, Management, and Security ...................................................................... 30 
11.3 Data Monitoring and Cleaning ....................................................................................... 30 


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Abbreviations 
CRSN: Centre de Recherche en Santé de Nouna 
DCC: Data Coordinating Center  
DSMC: Data and Safety Monitoring Committee 
GPS: global positioning system 
HDSS: Health and Demographic Surveillance Site 
IHPS: Infantile Hypertrophic Pyloric Stenosis 
IRB: Institutional Review Board 
MUAC: mid-upper arm circumference  
NP swabs: nasopharyngeal swabs 
PCR: polymerase chain reaction  
STGG: skim milk tryptone glucose glycerin media 
UCSF: University of California San Francisco  
WHO: World Health Organization 


Version 1.3 
5 
1. Chapter 1: Overview
1.1 Executive summary 
Although under-5 mortality rates are declining globally, neonatal mortality remains persistently high 
in many regions of sub-Saharan Africa.1 Mass azithromycin distribution to children aged 1-59 
months has been shown to reduce childhood mortality in Niger, Tanzania, and Malawi.2 This study 
did not evaluate the effect of azithromycin administered during the neonatal period. Observational 
evidence from high income countries has suggested that macrolides, including erythromycin and 
azithromycin, may be associated with increased risk of development of infantile hypertrophic pyloric 
stenosis (IHPS).3,4 However, these studies are limited by confounding by indication, as infants only 
receive antibiotics when they are ill.  
We proposed an individually randomized trial of azithromycin versus placebo to establish the 
efficacy and safety of administration of a dose of azithromycin during the neonatal period. Our 
long-term goal is to generate evidence that can be used by neonatal and child survival programs 
related to the use of azithromycin in the youngest children who have the highest risk of mortality. 
1.2 Objectives 
1: 
Establish the safety and efficacy of a single dose of azithromycin among neonates. 
Neonates aged 8 to 27 days will be randomized to a single dose of azithromycin or 
placebo. We hypothesize that neonates randomized to a single dose of azithromycin will have 
significantly lower all-cause mortality by 6 months of age, compared to those randomized to placebo. 
2:         Evaluate the impact of Neonatal azithromycin administration on long term 
microbiome and resistome in children enrolled in the NAITRE study two to five 
years after treatment administration. We hypothesize that neonates randomized to a single 
dose of azithromycin will have a significant difference in resistome and microbiome compared to 
children randomized to receive a single dose of placebo. 
1.3 Study Sites 
This study will be conducted in several regions of Burkina Faso, including peri-urban areas of 
Ouagadougou and Nouna town, and rural areas that are within 4 hours drive of a pediatric facility 
with capacity for performing pyloromyotomy.  


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2 
Chapter 2: Context 
Child mortality in West Africa is among the highest in the world. Although child health and 
mortality are improving worldwide, children in the Sahel and sub-Sahel regions of West Africa have 
the greatest risks of mortality.1,5 Burkina Faso’s current under-5 mortality rate is estimated 110 per 
1,000 live births5. Similar to other countries in the region, the major causes of child mortality in 
Burkina Faso are malaria, respiratory tract infection, and diarrhea. Malnutrition acts as a major 
underlying contributor to mortality.6,7 Neonatal mortality remains persistently high, with 
approximately 1/5th of neonatal mortality due to pneumonia, meningitis, and sepsis.8 Interventions 
that address these underlying causes may be particularly efficacious for reducing mortality. 
Younger children are at a higher risk of mortality. Approximately 2/3rd of under-5 deaths occur 
during the first year of life.5 In general, the child mortality rate decreases as age increases. While 
some improvement has been observed, neonatal mortality is declining at a slower rate than post-
neonatal childhood mortality.5 Many child health interventions are designed specifically for children 
over 6 months of age, such as vitamin A supplementation, seasonal malaria chemoprevention, and 
lipid-based nutritional supplementation. Identification of strategies that are safe and effective for the 
youngest children will be required to address persistently high rates of neonatal and infant mortality. 
The MORDOR I study demonstrated a significant reduction in all-cause child mortality 
following biannual mass azithromycin distribution. Across three diverse geographic locations in 
sub-Saharan Africa (Malawi, Niger, and Tanzania), biannual mass azithromycin distribution over a 
two-year period led to a 14% decrease in all-cause child mortality. In Niger, 1 in 5-6 deaths were 
averted. These results are qualitatively similar to those of a previous study of mass azithromycin 
distribution for trachoma control in Ethiopia, which found reduced odds of all-cause mortality in 
children in communities receiving mass azithromycin compared to control communities.9  
In MORDOR I, the strongest effect of azithromycin was in the youngest cohort of children. 
Across all three countries, the strongest effect of azithromycin was consistently in children 1-5 
months of age, with an approximately 25% reduction in all-cause mortality. However, MORDOR I 
was not optimized to target the youngest age groups. Although children as young as 1 month were 
eligible, biannual distributions might not reach some children until 7 months of age. On average, 
children were first treated at 4 months. Given that there may be a substantial benefit to treating 
children at younger ages, azithromycin strategies that are designed to target younger age groups may 
be even more beneficial for reducing child mortality.    
Here, we propose a randomized controlled trial designed to evaluate the efficacy of a dose of 
azithromycin administered during the neonatal period for prevention of mortality within in 
the first year of life. We propose to randomize births in several geographic regions of Burkina Faso 
to a single dose of azithromycin or placebo between day 8 and 27 of life. This study is designed to 
provide evidence of the efficacy of azithromycin treatment for the youngest children. 


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3 
Chapter 3: Study Design 
We will enroll and follow children from birth until 6 months of age. Figure 1 shows the trial profile. 
Figure 1. Trial profile 
Live Births Identified 
Assessed for Eligibility 
Randomized 
Allocation to Azithromycin 
Allocation to Placebo 
Treatment (Day 8 to 27) 
Treatment (Day 8 to 27) 
Treatment Day + 21 
Treatment Day + 21 
Day 90 
Day 90 
Day 180 
Primary Outcome: 6 month 
mortality 
Anthropometry  
Day 180 
Primary Outcome: 6-month 
mortality 
Anthropometry 
Day 365 
Infant Mortality Assessment 
Day 365 
Infant Mortality Assessment 
2 years after treatment 
Long term follow up 
2 years after treatment 
Long term follow up 
5 years after treatment 
Long term follow up 
5 years after treatment 
Long term follow up 
3.1 Recruitment 
Mothers and infants will be recruited either during antenatal care visits, via facility births, or via a key 
informant who notifies study staff of a birth. 


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3.1.1 
Antenatal clinic recruitment 
Women in the third trimester of their pregnancy who attend antenatal care services in health 
facilities in the geographic catchment areas of the study will be approached for possible inclusion in 
the study. Study staff members will approach women in late pregnancy and explain and discuss the 
study with the woman. If the woman is interested in participating, her contact information and 
approximate due date will be recorded by the study staff member. For women who are recruited 
during pregnancy, we will obtain verbal consent from the woman to follow the pregnancy until 
delivery. Once the baby is born we will obtain individual written consent to enroll the infant in the 
study. Formal eligibility assessment and enrollment will occur after birth. 
3.1.2 
Facility birth recruitment 
Newborns and mothers will be recruited in facilities that have delivery services. Postpartum women 
will be approached by study staff, who will explain the study to the woman and assess the newborn’s 
eligibility for the study. Newborns will be enrolled on the day of recruitment if they meet eligibility 
criteria (Chapter 4). 
3.1.3 
Key informant-based recruitment 
In some rural areas and in areas with Health and Demographic Surveillance Sites (HDSS) we will 
employ key informants who will alert study staff of new births in their catchment area. Key 
informants will receive a small remuneration for reporting births.  
3.2 Enrollment 
Enrollment will occur the same day the child will be treated. 
For each child assessed, a study staff member will review the eligibility criteria and fill out an 
eligibility assessment form (Appendix 2). If a child meets all of the eligibility criteria, a written 
informed consent process will be undertaken with the child’s caregiver (Chapter 9. After eligibility 
assessment and consent, the child will be formally enrolled in the trial, a study identification number 
will be assigned to the child, and the child will be randomized a treatment arm. 
3.3 Randomization 
3.3.1 
Unit of randomization  
The unit of randomization is the neonate. Children will be randomized to azithromycin or placebo. 
3.3.2 
Randomizing treatment 
Children will be randomized in a 1:1 fashion to a single dose of azithromycin or placebo. 
Randomization will occur via the tablet based on the child’s identification number. Children will be 
assigned a random letter that corresponds to the treatment bottle. To prevent accidental unmasking, 


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9 
a total of 8 letters will be used in the study, with 4 referring to azithromycin and 4 to placebo. Each 
facility will be stocked with medication bottles labeled with each of the 8 letters. 


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4 
Chapter 4: Eligibility 
4.1 Eligible Communities 
To be eligible for the trial, a community must meet the following criteria: 
1.
Within 4 hours of a facility that can provide services for pyloromyotomy (Ouagadougou or
Bobo Dioulasso)
2.
Accessible during the rainy season
3.
Ultrasound machine available OR a facility in which an ultrasound machine could be placed
is within 2 hours
4.2 Eligible Participants 
Eligible participants are neonates who are screened within the first week of life. Specific inclusion 
and exclusion criteria are described below. 
Inclusion Criteria (all must be met): 
•
Weight ³2500 g
•
Able to feed orally
•
Family intends to stay in study area for at least 6 months
•
Appropriate consent from at least one caregiver
•
No known allergy to azalides
•
Not living within one of the communities included in the community study
(CHAT/CHATON)
•
No hepatic failure manifested by neonatal jaundice
Exclusion Criteria (any excludes): 
•
Weight <2500 g
•
Unable to feed orally
•
Family planning to move
•
Mother/caregiver not willing to participate
•
Allergic to azalides
•
Living in one of the communities included in the community study (CHAT/CHATON)
•
Hepatic failure manifested by neonatal jaundice
Only children who meet all inclusion criteria and do not meet any exclusion criteria will be enrolled 
in the trial.  
Inclusion criteria for the long term follow up 


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11 
 
• Enrolled in the NAITRE study 
• Written consent of one guardian is obtained 
• Randomly selected to participate in the long term follow up  


 
12 
5 
Chapter 5: Procedures 
 
A general overview of study procedures is shown in Table 1. Eligibility assessment, enrollment, and 
randomization are covered in detail in Chapters 3 and 4. Appendix 2 contains all study forms. 
 
Table 1. Overview of study procedures 
 
 
Day (since birth) 
Activity 
Study Form 
0 - 7 
• Birth 
• Sensitization 
 
 
8 to 27  
• Assessment of eligibility  
• Enrollment 
• Randomization 
• Anthropometry (weight, 
Length, MUAC) 
• Treatment  
Form 1: Eligibility assessment 
Form 2: Baseline  
Form 3: Treatment 
Treatment Day + 21 
• Field worker safety assessment 
including vital status 
Form 4: Vital status 
Form 5: Neonate AE and 
IHPS risk 
90  
• Field worker safety assessment 
including vital status 
Form 4: Vital status 
Form 5: Neonate AE and 
IHPS risk 
180 (primary outcome) 
• 6 months vital status 
assessment 
• Anthropometry 
Form 6: 6 months vital status 
Form 
Form 5: Neonate AE and 
IHPS risk 
365 
• Vital status assessment 
Form 4: vital status 
2 years after treatment 
• Long-term follow-up in a 
randomly selected subgroup of 
children including, vital status 
assessment, survey on 
antibiotic use and biological 
sample collection 
 
Form 8: long term follow up 
form 
Form 9: long term survey 
5 years after treatment 
• Long-term follow-up in a 
randomly selected subgroup of 
children including, vital status 
assessment, survey on 
antibiotic use and biological 
sample collection 
•  
Form 8: long term follow up 
form 
Form 9: long term survey 
 
5.1 Treatment 


 
13 
 
Details on study medication are provided in Chapter 8. No child will be treated before Day 8 or after Day 27. 
After treatment, the study staff member will record the treatment or if the child did not receive the treatment, 
and for those who were not treated, the reason the child did not receive the treatment. 
 
5.2 Field worker safety assessment 
 
Field workers will conduct an initial home visit 21 days after treatment, followed by a home visit at 3 and 6 
months after birth. At 21 days following treatment, field workers will conduct a survey of adverse events, 
including if the child had any of the following symptoms: 
• Fever 
• Diarrhea 
• Vomiting 
• Abdominal pain 
• Skin rash 
• Constipation 
 
The field worker will assess whether the caregiver had sought care for the child since the last time they spoke 
to the study team, and if so, what the reason was for the health care visit and if the child was hospitalized.  
 
All caregivers who report that the child has been vomiting since the last study visit will be asked additional 
follow-up questions to screen for IHPS, including questions related to progression of vomiting, if the vomiting 
is projectile, and if the child is not gaining or losing weight. Those who respond affirmatively to those 
questions will be referred to the study pediatrician for additional follow-up and possible referral to a tertiary 
facility. (Appendix 3) 
 
All caregivers will be given a pamphlet explaining the signs of IHPS. We will also include contact information 
and instructions on what to do if the infant develops symptoms. (Appendix 4). 
 
5.3 Vital status assessment  
 
A vital status assessment will occur at each follow-up study assessment. Field workers will record on the tablet 
if the child is alive, died, moved, or unknown. 
 
5.4 Anthropometry assessment 
 
We will record every enrolled child’s weight and length at baseline and at 6 months of age. The anthropometry 
measurements will be taken at baseline when the neonate is enrolled and during the in person follow-up visit 
on day 180. Weight will be measured using a digital scale. Length will be taken using a Shorrboard to the 
nearest cm. We will also record the middle upper arm circumference of every enrolled child. (See appendix: 
anthropometric measurements protocol). 
 
5.5 Passive Surveillance 
 
In the Centre de Santé et de Promotion Sociale (CSPS, community health facility), we will conduct morbidity 
passive surveillance. Each CSPS will be equipped with a tablet for electronic capture of health facility visits. 
Each visit will be recorded, including the reason for the visit (e.g., fever, diarrhea, malnutrition, etc), the village 


 
14 
of residenc, the person’s age and sex, diagnosis (e.g., malaria, pneumonia, etc), treatment (e.g., antibiotic, 
antimalarial, etc), and timing of the visit (e.g., first versus follow-up visit). Note that this data is already 
routinely collected on paper forms. Identifying information like names will not be collected. 
 
 
5.6 Long term follow up  
 
Long term follow up will be performed on a randomly selected subgroup of children 2 years and 5 years after 
treatment in the same subset of children. 
The long term follow up will include a vital status check, a survey on antibiotic use, rectal sample, 
nasopharyngeal samples and blood will be collected to assess for antimicrobial resistance, microbiome diversity 
and blood transcriptome. 
 
Antibiotic survey 
The antibiotic survey will be performed by medical personnel previously trained during the long term follow 
up. Data will be recorded electronically using an mobile device with custom made application.  
Question of the survey will be asked to the caregiver of the child. The survey will ask whether the child have 
sought care to the health center or hospital since the beginning of the study and if they did it will ask if 
antibiotic were prescribed and what type. Pictures of most commonly used antibiotics will be used to help the 
caregiver answer the questions. 
 
Rectal swabbing  
Rectal swabbing will be performed during the long term follow up visits at 2 years and 5 years after treatment 
in a random subset of participants. The collection will be performed by medical personnel previously trained. 
Rectal swab will be collected in the following way: 
1. Put on a clean pair of gloves. 
2. Partially open the fecal swab package and remove the top section of the collection vial 
(this can be discarded). 
3. Position the child: 
• Lie the child on his/her back, hold legs in the air (it is useful to have assistance). 
• Or have the child lay on his/her stomach across the mother/guardian’s lap 
4. Remove the swab from the package. Take care that the cotton tip is not touched. If it is 
touched, throw the swab away and begin with a new one. 
5. Insert the tip of the swab into the child’s anus only as far as needed to contact fecal 
material (1-3cm) and rotate 180 degrees. The tip should be a brownish color when 
removed. 
6. Place swab into the preservative in the collection tube. Make sure the swab is fully 
submerged in the liquid preservative and then break the swab off using the pre-scored 
breaking point. 
7. Screw the cap back on the tube and make sure that it’s tightened. Wrap the area where the 
cap meets the tube with Parafilm to ensure that the sample will not leak, and then place 
the tube into the appropriate sample box. 
• If the swab cannot be broken off while the tip is fully submerged in the liquid, try 
twirling the swab in the liquid first (to release the contents of the sample into the 
preservative) before breaking it off. Avoid rubbing the sample on the tip of the swab 


 
15 
off on the side of the tube where there is no liquid. 
8. Place a random number label on the collection tube. 
9. Place the tube the rectal swab container. 
10. Swab storage for Genetic analysis: Store samples at room temperature. According to the 
manufacturer, the preservative in the tube will preserve DNA for 5 months at room 
temperature. Samples can be frozen for long time storage 
 
 
 
Nasopharyngeal swabbing 
 
Nasopharyngeal swabbing will be performed during the long term follow up visits at 2 years and 5 years after 
treatment in a random subset of participants. The collection will be performed by medical personnel previously 
trained and will be collected in the following way: 
 
The examiner will: 
 
1. 
Place a pediatric flocked swab with a nylon tip through the right nostril and down the 
nasopharynx of each participant. Note that if the swab is not perpendicular to the frontal 
plane of the face, it is likely not in the inferior turbinate. 
 
2. 
Reach the nasopharynx, rotate the swab 180° and remove the swab from the nose. 
 
3. 
Place the swab in a tube containing 1.0 mL DNA/RNA shield media by Zymo media, cut 
the handle off using sterile scissors, and close the cap of the tube with the swab immersed. 
 
4. 
The nasopharyngeal swab samples in DNA/RNA shield media will be stored in ambient 
temperature in the field. Then transferred to a refrigerator or freezer. 
 
5. 
The scissors used to cut calcium alginate swabs will be sterilized with alcohol pads or 
cleaned with bleach wipes between participants. When collecting specimens in 
DNA/RNA shield, scissors will be cleaned between participants - first with bleach 
wipes, and then with alcohol pads. 
 
Do not attempt to collect the NP swab if you are not successful after three attempts. 


 
16 
Nasopharyngeal swabs will be stored in DNA/RNA shield media by Zymo research, and will be 
assessed for the most common genetic resistant determinants (ermB and mefA) using a PCR-based 
assay. Serotype will be assessed using a nested PCR reaction for the most common serotypes, 
followed by the Quellung reaction for any untyped isolates. 
 
Blood collection 
Blood collection will be performed during the long term follow up visits at 2 years and 5 years after treatment 
in a random subset of participants. The collection will be performed by medical personnel previously trained 
and will be collected in the following way: 
 
Determination of the maximum allowable blood draw volume 
See the chart below in order to determine the maximum allowable blood draw volume from children 
and infants, based on body weight. 
 
Body wt in kg 
Max drawn in one blood 
draw 
 
2.5% of total blood volume 
1 kg 
2.5 ml 
2 kg 
4.5 ml 
3 kg 
6 ml 
4 kg 
8 ml 
5 kg 
10 ml 
6 kg 
12 ml 
7 kg 
14 ml 
8 kg 
16 ml 


Version 1.3 
 
17 
9 kg 
18 ml 
10 kg 
20 ml 
11 thru 15 kg 
22 - 30 ml 
16 thru 20 kg 
32 - 40 ml 
21 thru 25 kg 
42 - 50 ml 
26 thru 30 kg 
52 - 60 ml 
31 thru 35 kg 
62 - 70 ml 
36 thru 40 kg 
72 - 80 ml 
41 thru 45 kg 
82 - 90 ml 
46 thru 50 kg 
92 - 100 ml 
Greater than 50 kg 
100 ml 
 
Citation for above chart: Seattle Children’s Hospital Powered by Mayo Medical 
Laboratories. “Maximum Allowable Blood Draw from Infants.” 
https://seattlechildrenslab.testcatalog.org/show/1000721-1 
 
Identification du patient: 
Identify the child by scanning her/his study identity card. 
Verify the identity of the child with the mother/guardian of the child: check the name, study 
identification number, sex, age and the name of the mother and father of the child. 
 
Supplies  
2 chairs 
1 examination table 
1 pillow 
1 Cotton ball 
1 Alcohol wipes 
1 Wing needle 23G with tubing and holder 1 needle container 
1 trash bag 
1 small bandage 1 pair of gloves 
1 tube to collect blood  
1 tourniquet 
1 tablet 
1 unique identification number 
1 blue pad to put under the arm of the child 
 
Procedure 


Version 1.3 
 
18 
1. 
Collect supplies and equipment 
2. 
Wash your hands 
3. 
Look at the arms of the child to try to find a good venipuncture site 
4. 
Wash your hands 
5. 
Position the child 
6. 
Put the tourniquet on the child about 2 fingers widths above the venipuncture site chosen 
7. 
Put on your gloves 
8. 
Attach the end of the tubing to the end of the holder 
9. 
Remove the plastic sleeve from the needle 
10. 
Using the alcohol swab, disinfect the venipuncture site and allow it to dry 
11. 
Draw the skin using your tumb and stick the needle in the vein 
12. 
Push the tube onto the holder until the needle is in it 
13. 
Blood should begin to flow into the tube 
14. 
Fill the tube until it is full, remove the tube 
15. 
Release the tourniquet 
16. 
Place a dry coton ball over the venipuncture site and withdraw the needle 
17. 
Ask the parent/guardian to continue applying mild pressure 
18.       Collect dried blood spots using the whole blood 
19. 
Dispose of the needle in the sharps container 
20. 
Dispose of all contaminated supplies in the trash 
21. 
Label the tube with a label containing a unique number 
22. 
Put a bandage on the child’s arm and ask the mom to remove it 1h later 
23. 
Gently and completely invert the tube 10 times in order ro mix blood with the tube 
preservation agents 
24. 
Remove gloves, put them in the trash 
25. 
Wash your hands 
 
How to fill out the filter paper: 
1. Label the filter paper with a random number sticker.  
2. Grip the filter paper on the side without small circles. Place a droplet of blood directly 
from the thumb or finger onto five of the six circles, leaving the right one blank. Be sure 
to fill each circle completely.  


Version 1.3 
 
19 
 
 
 
 
 
 
 
 
 
 
 
 
3. The recorder will scan the QR code. 
4. Carefully slide the filter paper onto a pencil to air dry for at least an hour. There should 
be about 1 cm in between each sample. Secure the pencil into a Styrofoam surface in a 
box or container to protect from dust. 
5. When the filter paper is dry, place each sample into a small zip plastic bag (individually). 
Place the small bags into a larger Ziploc bag with five desiccant packets.  
6. Ensure the large Ziploc bag is sealed tightly, as moisture will damage the samples. 
Transport these filter paper samples to a freezer. 
 
If venipuncture is not possible at the study site, we will collect blood using fingerprick method. 
 
6 
Chapter 6: Adverse Event Monitoring and Safety Assessment  
 
There is some concern that azithromycin administered during the neonatal period will increase the 
risk of infantile hypertrophic pyloric stenosis (IHPS). Follow-up assessments have been designed to 
carefully monitor for IHPS following administration of study treatment. 
 
6.1 Background on Infantile Hypertrophic Pyloric Stenosis in Neonates 
 
IHPS is a condition in which the pylorus of the stomach becomes thickened, resulting in gastric 
outlet obstruction.10 In developed countries, the incidence of IHPS is approximately 2 per 1,000 
infants in the general population. While surgery is generally curative, the condition is lethal in the 
absence of surgery. The cause of IHPS is unknown and likely complex, and both genetic11,12 and 
environmental13,14 factors are thought to contribute to its development. Male neonates are 
disproportionately affected by IHPS, with a 4-5:1 male to female ratio.10,12,15 Other risk factors for 
IHPS include prone sleeping position, bottle/formula feeding,14,16,17 preterm birth,13,14 cesarean 
section delivery,13,14 and birth order.10,13-15 IHPS incidence appears to be decreasing over time, which 
some have attributed to public health interventions promoting supine sleeping position and 
exclusive breastfeeding.18,19 
 
In addition to genetic and environmental factors, erythromycin is associated with increased risk of 
infantile hypertrophic pyloric stenosis (IHPS).20 Given that azithromycin is a related compound, 
there is some concern that azithromycin may lead to increased risk of IHPS in neonates. Limited 
Area to hold the filter paper.  
Do not touch the small circles. 
Leave last circle blank 


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20 
evidence exists of the risk of IHPS among neonates treated with azithromycin.21 Two randomized 
controlled trials have assessed the use of intravenous azithromycin for prevention of 
bronchopulmonary dysplasia (BPD) in low birthweight infants (<1,250g) compared to placebo 
within 72 hours of birth.22,23 Of 263 neonates enrolled in the two studies (N=130 receiving IV 
azithromycin), no cases of IHPS were reported. A third non-placebo controlled randomized trial of 
azithromycin prophylaxis for BPD in premature neonates additionally reported no cases of IHPS 
among 53 neonates receiving azithromycin.24 An observational study of 58 infants receiving 
azithromycin following exposure to a healthcare worker with pertussis did not identify any cases of 
IHPS.25 The largest study of azithromycin exposure in neonates is a retrospective cohort of more 
than one million infants in the TRICARE Management Activity military health system, which 
reported an overall IHPS rate of 2.3 per 1,000 (95% CI 2.2 to 2.4) among children in the first 90 
days of life.4 Of 4,875 infants prescribed azithromycin, there were 8 cases IHPS, 3 of which 
occurred when azithromycin was prescribed during the first 14 days of life and 5 of which during the 
first 15-42 days of life. Overall, there was no significant difference in IHPS rate in infants treated 
with azithromycin versus IHPS rates among infants who had not received an antibiotic during the 
first 90 days of life were not presented by time since birth. Table 2 lists the rates of IHPS following 
azithromycin, erythromycin, or cephalexin prescription.  
 
Table 3. Unadjusted rate of IHPS in infants with oral azithromycin, per 1,000 
 
Azithromycin 
Erythromycin 
Cephalexin 
0-14 days1 
15-42 days1 
43-90 days1 
Overall 
20.3 (4.2-59.2) 
6.9 (2.2-16.1) 
0 (0-0.9) 
1.6 (0.7-3.2) 
30.9 (14.1-58.7) 
9.3 (3.0-21.8) 
2.8 (0.6-8.1) 
8.9 (5.2-14.3) 
2.2 (0.05-
12.2) 
3.1 (1.0-7.2) 
0 (0-1.6) 
1.4 (0.5-3.0) 
1Day of azithromycin prescription, from birth 
 
There is relatively little evidence of the epidemiology of IHPS in sub-Saharan Africa. A study in 
Nigeria documented 57 cases of IHPS from 1978-2008 at a university teaching hospital, with only a 
single case from 2003-2008.19 A study of 102 cases of IHPS over a 5-year period in Tanzania 
documented a 4.9% mortality rate despite surgical intervention.26 The risk of mortality was higher in 
infants under 2 weeks of age and those with delayed presentation to care. A retrospective study at a 
tertiary hospital in Ethiopia found that 12.9 per 1,000 admissions were due to IHPS, with a 3.3% 
mortality rate.15 In Ouagadougou, an unpublished case series of 32 infants treated for IHPS showed 
a 6.3% mortality rate, which was attributed to late presentation due to the infant traveling from far 
outside the city. 
 
6.2 Screening for IHPS 
 
Field workers will conduct home visits with caregivers 21 days following treatment, as described in 
Chapter 5. Any child suspected of having IHPS will be immediately referred to the study pediatrician 
for evaluation. Evaluation will include ultrasonography and physical exam. Physical exam will 
include assessment of the pyloric olive (a thickened and elongated pylorus). Any child with a positive 
physical exam for a pyloric olive will be immediately transferred to the pediatric surgical unit in 
Ouagadougou or Bobo. 
 


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21 
Images will be taken including the longitudinal pylorus with canal length measurement, transverse 
pylorus with muscle thickness measurement, and the relationship of the pylorus to the gallbladder. 
IHPS will be strongly suspected in infants with a permanently closed pylorus and exaggerated, 
retrograde gastric peristalsis. Diagnostic measurements include pyloric muscle thickness (diameter of 
a single muscular wall on a transverse image) >4 mm, length (longitudinal measurement) >15 mm, 
and pyloric volume >1.5 cc.27 The child should be placed with their right side down, and the pylorus 
watched to determine if it opens. Small infants not below the pathologic limits with a permanently 
closed pylorus will also be considered for further workup.28 Any child with a pyloric muscle 
thickness >4 mm will be immediately transferred to the pediatric surgical unit in Ouagadougou. In 
addition any child with normal measurements, but no food is passing, will be transferred.  
 
All children will be assessed for electrolyte disturbance and dehydration, and rehydration and 
correction of electrolyte disturbance will occur prior to surgery. 
 
Any children receiving surgery for IHPS will be followed-up with 1 week after the procedure. The 
follow-up will be a phone call or at-home visit. The child’s vital status information will be collected. 
Another follow-up visit will take place 4 weeks after the procedure.  
 


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22 
7 
Chapter 7: Training  
 
CRSN and UCSF will work together prior to the start of the study to standardize all study 
procedures. We will review the format, general logistics, and procedures for the recruitment, 
enrollment, randomization and anthropometric measurements. We will review all study protocols, 
including informed consent procedures and documentation and adverse event monitoring. The 
importance of screening for IHPS and safety protocols will be stressed. Refresher trainings will 
occur as needed and supervisors and study investigators will routinely review data and procedures to 
ensure fidelity to protocol.
8 
Chapter 8: Study Medication 
 
Neonates enrolled in the study will be offered weight-based, directly observed, oral suspension 
azithromycin or placebo. We will monitor adverse events following treatment as described in 
Chapter 6.  
 
8.1 Study Medication Description (from Pfizer, Inc.) 
 
Azithromycin 
 
Zithromax® for oral suspension is supplied in bottles containing azithromycin dehydrate powder 
equivalent to 1200mg per bottle and the following inactive ingredients: sucrose; tribasic  anhydrous 
sodium phosphate; hydroxypropyl cellulose; xanthan gum; FD&C Red #40; and flavoring including 
spray dried artificial cherry, crème de vanilla, and banana.  After constitution, a 5mL suspension 
contains 200mg of azithromycin. 
 
8.2 Dosage Information 
 
Azithromycin and placebo will be administered as a single dose, in oral suspension form for 
children. Dosing will follow the WHO recommendations for treatment of active trachoma: 
• Single dose of 20mg/kg in children (up to the maximum adult dose of 1g)  
 
Individuals who are allergic to macrolides/azalides will not be treated. 
 
8.3 Medication Procurement/Donation 
 
Azithromycin (Zithromax®) and the placebo have been donated by the Pfizer Corporation.  There 
will be no costs to acquiring the study medication. Pfizer, Inc. will ship azithromycin and placebo 
directly to the study sites. Representatives of each study site will manage the customs process and 
transport the medication from the port to storage sites.   
 
8.4 Medication Quality Control 
 


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23 
Study medication will be shipped by Pfizer directly to CRSN and stored at CRSN research offices 
prior to distribution to each study site.  The study coordinator and other staff will regularly check 
and record the study medication expiration dates.  The expiration dates on the medication 
containers will be strictly monitored and all expired study medicine will be discarded appropriately. 
The study coordinator will work with each health facility to ensure that they have appropriate stock 
of all study medications. 
 
8.5 Study Treatment 
 
Study treatment procedures are detailed in Chapter 5.  
 
8.6 Serious Adverse Events 
 
Any serious adverse events (SAE) will be reported to Pfizer. An IIR SAE Form (Investigator-Initiated 
Research Serious Adverse Events Form) will be completed for each event. (See Appendix for form and 
complete instructions.) 
 
According to Pfizer, an SAE is any adverse event that: 
• Results in death 
• Is life-threatening (i.e., causes an immediate risk of death) 
• Requires inpatient hospitalization or prolongation of existing hospitalization 
• Results in persistent or significant disability or incapacity  
• Results in a congenital anomaly or birth defect 
 
 
Or that is considered to be: 
 
• An important medical event 
 
All participants will be advised to alert a village health worker if they experience, within one week of 
treatment, a serious adverse event (by the preceding definition). An SAE report must be submitted 
for all deaths in the study – regardless of the time of treatment. The local health worker will report 
to the study coordinator; who must, within 24 hours, submit a Pfizer IIR SAE Form to 
mordor.burkina.sae@gmail.com. AS and TL will review, and forward to Pfizer and/or the Medical 
Monitor, as appropriate. SAEs must be submitted to Pfizer within 24 hours of receipt from the on-
site coordinator. AS and TL will also forward SAE to DSMC if meets criteria of being possibly 
related to study drug. All deaths reported that are not serious adverse events resulting from the 
treatment will be reported to Pfizer on a quarterly basis.  
 
One or more qualified investigators will be posted in the health centers at the recruitment site. 
Mothers of included children will be encouraged to visit these health centers in the event of an 
adverse event. Any adverse events occurring during the trial will be covered by the study free of 
charge. Participants with a serious event will be cared for in the nearest hospital. The research team 
will be in constant contact with pediatricians from regional and university hospitals and pediatric 
surgeons to manage potential serious adverse events. Surgical units will be involved in the diligent 
management of pyloric stenosis cases. All recruitment sites will be within a two-hour drive of a 


Version 1.3 
 
24 
Regional Hospital Centre where there is a paediatrician and a radiologist to enable rapid diagnosis of 
pyloric stenosis and within 4 hours of Bobo-Dioulasso or Ouagadougou to enable rapid 
management.  
 
The reporting of any serious adverse event will follow national procedures in Burkina Faso. 
- In the event of a non-serious adverse event, the CSPS will process the cases and the national 
reporting form will be completed. 
- In the event of serious events, the CSPS will contact the study doctors on the same day. The 
patient will then be evacuated to an appropriate level of care for management. 
- The declaration will be made to the National Agency for Pharmaceutical Regulation in accordance 
with the regulations and within the deadlines (7 days in the event of death or life-threatening 
prognosis, 15 days in other serious and unexpected cases, 15 days in new facts) at 
pharmacovigilance.burkina@sante.gov.bf  
- The ethics committees will also be informed of the occurrence of this event within the same time 
frame. 
- The sponsor will be informed within 24 hours in the event of a serious event. 
Pfizer will be reported within 24 business hours from the date of knowledge of the serious adverse 
event 
 
8.7 Adverse Events Data 
We will keep records and report all adverse events of azithromycin to the DSMC. We will report 
both efficacy and side effects of azithromycin For any “sudden deaths” believed to be associated 
with azithromycin treatment, key informants will immediately notify the verbal autopsy interviewer 
via SMS message or another appropriate form of rapid communication.


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25 
9 
Chapter 9: Protection of Human Subjects 
 
Before the study begins, CRSN and UCSF will obtain formal ethical approval from their respective 
ethics committees as well as national ethical approval in Burkina Faso. In addition, local staff will 
approach community leaders to describe the study and answer any questions. Study staff will 
proceed only if local leadership consents to participate. At the individual level, we will obtain written 
consent from a parent or guardian for all study activities with patient contact, including following 
pregnancies for potential enrollment, treatment, and follow-up visits. 
 
If, at any time, a parent or guardian elects to withdraw a family member from the study, they will be 
free to do so. Individuals who withdraw will be offered the same medical treatment outside the 
study. 
 
 
9.1 Institutional Review Board Approval 
 
UCSF Committee on Human Research 
UCSF’s Committee on Human Research will annually review study protocol for ethical approval. 
 
CRSN Comité Institutionnel d’Ethique  
The study protocol will be reviewed and granted ethical approval by the Comité Institutionnel 
d’Ethique at the CRSN headquarters before any patient-related research activities begin. 
 
National Health Ethics Committee of Burkina Faso. 
The study protocol will be reviewed and granted ethical approval by the National Health Ethics 
Committee of Burkina Faso before any patient-related research activities begin and annually. 
 
9.2 Informed Consent 
First, the chairman of each village will be asked for permission to include the village in the study.  
Additionally, the study will be discussed with all adults in the village by team members who speak 
the local language(s). 
 
 
Informed consent scripts will be translated into local languages before the study can begin. Consent 
scripts will then be back-translated by a different party to ensure comprehension. Consent scripts 
will be submitted and approved by national IRB committees in Burkina Faso prior to study 
implementation. Then they will be read aloud to each study participant (and his/her 
parent/guardian) by a local team member who is a native speaker of the local language to ensure 
that they understand the risks and benefits of participating in all study activities. Young adults and 
children under 18 years of age, who cannot give consent by law, will be included in the study only 
following the receipt of verbal informed consent from a parent or guardian. If, at any time, a parent 
or guardian elects to withdraw themselves or a family member from the study it will be made clear 
that they can, without consequences. 
 
   
 


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26 
10 Chapter 10: Data and Safety Monitoring Committee Charter 
 
This Charter is for the Data Safety and Monitoring Committee (DSMC) for Neonatal Azithromycin to 
Prevent Infant Mortality. (OPP 1187628) 
 
The Charter will define the primary responsibilities of the DSMC, its relationship with other trial 
components, its membership, and the purpose and timing of its meetings. The Charter will also 
provide the procedures for ensuring confidentiality and communication, statistical monitoring 
guidelines to be implemented by the DSMC, and an outline of the content of the Open and Closed 
Reports that will be provided to the DSMC. 
 
10.1 Primary Responsibilities of the DSMC 
The DSMC will be responsible for safeguarding the interests of trial participants, assessing the safety 
and efficacy of the interventions during the trial, and monitoring the overall conduct of the trial. The 
DSMC will provide recommendations about stopping or continuing the trial. To contribute to the 
integrity of the trial, the DSMC may also formulate recommendations relating to the 
selection/recruitment/retention of participants, to protocol-specified regimens, and the procedures 
for data management and quality control.  
 
The DSMC will be advisory to the trial leadership group, hereafter referred to as the Steering 
Committee (SC). The SC will be responsible for promptly reviewing the DSMC recommendations 
and determining, whether to continue or terminate the trial, and to determine whether amendments 
to the protocol are required. If needed, the DSMC may seek the advice of a content expert outside 
of the committee.  
 
 


Version 1.3 
 
27 
10.2 DSMC Membership 
The DSMC is an independent multidisciplinary group consisting of epidemiologists, biostatisticians, 
bioethicists, and clinicians that collectively has experience in pediatrics, the management of 
infectious diseases, and in the conduct and monitoring of randomized clinical trials including 
subsaharan Africa. 
 
10.3 Conflicts of Interest 
The DSMC membership has been restricted to individuals free of apparent conflicts of interest. The 
source of these conflicts may be financial, scientific, or regulatory. Thus, neither study investigators 
nor individuals employed by the sponsor, nor individuals who might have regulatory responsibilities 
for the trial products, are members of the DSMC. 
 
The DSMC members will disclose to fellow members any consulting agreements or financial 
interests they have with the sponsor of the trial, with the contract research organizations (CRO) , or 
with other sponsors having products that are being evaluated or that are competitive with those in 
the trial. The DSMC will be responsible for deciding whether these consulting agreements or 
financial interests materially impact their objectivity.  
 
The DSMC members will be responsible for advising fellow members of any changes in any of the 
membership requirements that occur during the course of the trial. It may be appropriate for DSMC 
members who develop significant conflicts of interest resign from the DSMC.  
 
DSMC membership is to be for the full duration of the trial. If any members leave the DSMC, the 
SC, in consultation with the DSMC, will promptly appoint a replacement.   
 
10.4 Timing and Purpose of the DSMC Meetings  
 
Organizational Meeting 
The initial meeting of the DSMC will be an Organizational Meeting. This is during the final stages of 
protocol development and the purpose is to provide advisory review of scientific and ethical issues 
relating to study design to discuss the standard operating procedures and to discuss the format and 
content of the Open and Closed Reports that will be used to present trial results. 
 
The Organizational Meeting will be attended by all DSMC members, lead trial investigators, and the 
trial biostatistician. The DSMC will be given the drafts of the trial protocol, the Statistical Analysis 
Plan, the DSMC Charter, and the current version of the case report forms. At subsequent meetings, 
committee members will receive Open and Closed Data Reports. 
 
Formal Interim Analysis Meetings 
One or more ‘Formal Interim Analysis’ meetings will be held to review data relating to treatment 
safety and efficacy, and quality of trial conduct. There will be at least two interim decisions to be 
made by the DSMC, at approximately 12 months and 24 months into the study. 
 
10.5 Procedures to Ensure Confidentiality and Proper Communication 
To enhance the integrity and credibility of the trial, procedures will be implemented to ensure the 
DSMC has access to all emerging information from the trial regarding comparative results of 
efficacy and safety, aggregated by treatment arm.  


Version 1.3 
 
28 
 
Closed Sessions 
Sessions involving only DSMC members and, where appropriate, those unmasked trial investigators 
(on the Data Coordinating Committee) who generate the Closed Reports (called Closed Sessions) 
will be held to allow discussion of confidential data from the trial, including information about the 
relative efficacy and safety of interventions.  
 
At a final Closed Session, the DSMC will develop a consensus on its list of recommendations, 
including that relating to whether the trial should continue. 
 
Open Session  
In order for the DSMC to have access to information provided, by study investigators, or members 
of regulatory authorities, a joint session between these individuals and DSMC members will be held 
between the Closed Sessions.  
 
Open and Closed Reports 
For each DSMC meeting, Open and Closed Reports will be provided. Open Reports, will include 
data on recruitment and baseline characteristics, pooled data on eligibility violations, and 
completeness of follow-up and compliance. The study statistician (TCP) will prepare these Open 
Reports.  
 
Closed reports, available only to those attending the Closed Sessions of the meeting, will include 
analyses of primary and secondary efficacy endpoints, including subgroup and adjusted analyses, 
AEs and symptom severity, , and Open Report analyses that are displayed by intervention group. 
These Closed Reports will be prepared by the study biostatistician.  
 
The Open and Closed Reports should provide information that is accurate, with follow-up that is 
complete to within two months of the date of the DSMC meeting. The Reports should be provided 
to DSMC members approximately three days prior to the date of the meeting. 
 
Minutes of the DSMC Meeting 
The research team will prepare minutes for the open portion of the meeting, including the DSMC’s 
recommendations. 
 
Recommendations to the Steering Committee (SC) 
At each meeting of the DSMC during the trial, the committee will make a recommendation to the 
Steering Committee to continue or terminate. This recommendation will be based primarily on 
safety and efficacy considerations and will be guided by statistical monitoring guidelines defined in 
this Charter. 
 
Recommendations to amend the protocol or conduct of the study made by the DSMC will be 
considered and accepted or rejected by the SC. The SC will be responsible for deciding whether to 
continue or to stop the trial based on the DSMC recommendations.  
 
The DSMC will be notified of all changes to the protocol or to study conduct. The DSMC 
concurrence will be sought on all substantive recommendations or changes to the protocol or study 
conduct prior to implementation.  
 


Version 1.3 
 
29 
The SC may communicate information in the Open Report to the sponsor and may inform them of 
the DSMC recommended alterations to study conduct or early trial termination in instances in which 
the SC has reached a final decision agreeing with the recommendation. The SC will maintain 
confidentiality of all information it receives other than that contained in the Open Reports until after 
the trial is completed or until a decision for early termination has been made. 
 
10.6 Statistical Monitoring Guidelines 
The SC will propose statistical rules for a futility stopping rule (requested by the sponsor) and an 
efficacy stopping rule at the first DSMC meeting. A decision will be made whether the efficacy 
stopping rule is appropriate for the study. 
 
 


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30 
11 Chapter 11: Data Collection, Management, and Security 
 
11.1 Scope of Data 
Mortality and morbidity data will be collected in this trial. Data include treatment, vital status, 
adverse events, demographic and birth-related characteristics, and specimen collections. 
 
Demographic and Birth Data 
At baseline, trained field workers will collect information on the child’s gestational age and 
birthweight and basic demographic information about the mother (age, education, etc). 
 
Treatment Data 
Trained field workers who administer treatment will record each treatment dose in the mobile 
application at the time of treatment of the child. Data will include the child’s weight (used to 
calculate the dose), the dose, if the child received the treatment, and if not, why the child did not 
receive treatment. 
 
Vital Status Data 
Trained field workers will collect information on the vital status of each infant enrolled in the study 
at multiple pre-specified time points during the study. Vital status will include alive, died, moved, or 
unknown. 
 
Adverse Events 
Trained health workers will collect data on adverse events that arise during the study, including 
gastrointestinal symptoms, hospitalizations, and other healthcare visits.  
 
 
11.2 Data Storage, Management, and Security 
Data will be recorded electronically using handheld mobile devices with custom-made software 
applications and uploaded daily onto a secure, password protected, central server. Rapid transfer of 
electronically captured data will allow nearly real-time monitoring of activity at the study site. Each 
study site will have a local data coordinating center within the study area. All handheld devices and 
data entry coordinating centers will be password protected, and all changes in data will be noted, 
including the date of the change, and the person who made the change. To ensure the quality of the 
data, we will conduct training sessions before each biannual census where needed. The central 
database application will use hard disk encryption and physical protection of the server (which is to 
be maintained in a locked room accessible only to authorized personnel). The database will be based 
on mySQL (which supports standard SQL queries). Data will be backed up off site (providing 
integrity in case of the physical loss of the server). Data will never be deleted from mobile capture 
devices until at least one offsite backup has been completed. Data security during electronic transfer 
will be achieved through use of the Advanced Encryption Standard (AES). 
 
11.3 Data Monitoring and Cleaning  
Data monitoring and cleaning will be overseen by the data coordinating center (DCC) at the 
coordinating site. Data collection will be monitored on a weekly basis by the site study coordinator 


Version 1.3 
 
31 
using a dashboard function. The dashboard will consist of the following reports by study site: Date 
Household Census Completed, Number of Households Census Completed by Village, Percent 
Household Census Completed by village, Treatment Status by Worker, Age Distribution by Worker, 
Sex Distribution by Worker, GPS Missing by Worker, GPS Missing by Village, Number of Records 
Synced by Date, Assigned Treatment by Given Treatment, Treatment Status by Age, Treatment 
Status by Village, Age Distribution by Village, and Sex Distribution by Village. 
 
The DCC will ensure that the site study coordinators log on to the dashboard at least weekly to 
confirm the status of the dashboard. In addition, upon each village census completion, the DCC will 
create and maintain a Stata program to identify data quality concerns. Any such concerns, which 
must be addressed at the site-specific level, will be queried by the DCC. At every phase, as each 
village is completed and the data is considered cleaned, the data will be locked and a list of deaths 
will be generated and provided to each site for verbal autopsy.  
 


Version 1.3 
 
32 
 
Appendix 
 
Appendix 1. Revision History 
Appendix 2. Study Forms 
Appendix 3. Referral decision tree 
Appendix 4. IHPS Pamphlet  
Appendix 5. Anthropometry protocol 


Version 1.3 
 
33 
References  
 
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Krogh C, Biggar RJ, Fischer TK, Lindholm M, Wohlfahrt J, Melbye M. Bottle-feeding and the 
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SAP_001.pdf: 
 
 
1 
 
 
NAITRE 
MORDOR Project 
June 10, 2021 
Version 2.1 
 
 
 
 
 
Statistical Analysis Plan 
 
 
 
 
 
UCSF Francis I. Proctor Foundation 
Centre de Recherche en Sante de 
Nouna University of Heidelberg 


 
 
2 
 
Introduction 
 
This document (Statistical Analysis Plan, SAP) describes the planned analysis and 
reporting for the clinical trial, Azithromycin for the prevention of neonatal mortality in 
Burkina Faso: an individual-randomized trial. It includes specifications for the 
statistical analyses and tables to be prepared for the interim and final Clinical Study 
Report.  This study is a Phase IV clinical trial to compare methods to reduce childhood 
mortality using mass administration of azithromycin (Pfizer, CAS 83905-01-5) compared 
to placebo. The content of this Statistical Analysis Plan meets the requirements stated by 
the US Food and Drug Administration and conforms to the American Statistical 
Association’s Ethical Guidelines. 
The following documents were reviewed in preparation of this Statistical Analysis Plan: 
 
• Azithromycin for the prevention of neonatal mortality in Burkina Faso: an individual- 
randomized trial, Manual of Operations 
• Azithromycin for the Prevention of Neonatal, Infant, and Child Mortality in Burkina 
Faso and Safety Assessment, Proposal 
• Statistical Analysis Plan, MORDOR Trial 
 
• ICH Guidance on Statistical Principles for Clinical Trials 
 
The planned analyses described in this SAP will be included in future manuscripts. 
Exploratory analyses not necessarily identified in this Statistical Analysis Plan may be 
performed to support the analysis. Unplanned analyses not delineated in this Statistical 
Analysis Plan will be documented as such in the final Clinical Study Report and 
manuscripts. Note that unplanned analyses will be broadly grouped into two categories: 
1. Sensitivity analyses for pre-specified outcomes 
 
2. Other unspecified analyses 


 
 
3 
 
Sensitivity analyses are defined as alternate ways of modeling the primary outcome to 
ensure the finding was not dependent on the analytic plan, and these will not be subject 
to a multiple comparisons correction. Other unspecified analyses will be declared 
hypothesis generating or subject to an alpha level of 0.001. 
This document will be reviewed prior to the enrollment of patients. All subsequent 
changes will be indicated by detailed change log in the Appendix. 


 
 
4 
 
Contents 
1 Summary 
5 
1.1 
Mortality Trial 
5 
2 Statistical Analysis 
5 
2.1 
Planned Analyses 
5 
2.1.1 
Mortality Trial 
5 
2.2 
Interim Monitoring 
9 
3 Sample Size Considerations 
10 
4 Randomization 
12 
5 Abbreviations 
13 
6 Revision History 
15 
 


 
 
 
1 
Summary 
1.1 
Mortality Trial 
 
The trial profile is given in the Manual of Operations. In the mortality trial, infants are 
randomized to two arms: Azithro and Control. The trial is a placebo-controlled, double-
masked (i.e., double-blind), individual-randomized clinical trial. 
Objective. Establish the safety and efficacy of a single dose of azithromycin among 
neonates. Neonates age 8 to 27 days will be randomized to a single dose of azithromycin 
or placebo. We hypothesize that neonates randomized to a single dose of azithromycin 
will have significantly lower all-cause mortality by 6 months of age, compared to those 
randomized to placebo. 
 
2 Statistical Analysis 
2.1 Planned Analyses 
2.1.1 
Mortality Trial 
 
Children between the ages of 8 and 27 days may be enrolled in the trial and be offered 
placebo or azithromycin in a masked fashion. Any child known to be living at a six month 
follow-up visit is counted as alive, any child known to have died is counted as dead, and 
any other child is counted as missing or lost to follow-up. Children followed up within six 
weeks of the actual six month follow-up date will be included in primary analysis. Deaths 
that occur on the day of enrollment after randomization will be assigned 0.5 days of 
person-time. Deaths that occur between two follow-up timepoints will be assigned person-
time in accordance to the midpoint between the two follow-up dates (i.e. a death occurring 
between the 3-month and 6-month timepoint will contribute 1.5 months of additional 
person-time). As a sensitivity analysis, children that were followed up within twelve weeks 
of six month follow-up date will be analyzed. 


 
 
6 
 
Primary Analysis. 
The primary analysis will be conducted as binomial regression using complementary 
log-log link. Firth correction will be employed to ensure numerical stability.  Inference will 
be based on the permutation test. 
The analysis will be two-sided, with a type I error rate (alpha) of 0.05. 
Statistical considerations. 
 
• Statistical tests will be conducted with Monte Carlo permutation based on the 
randomization unit. The number of replications will be 10,000, unless the Monte 
Carlo confidence interval for the P-value includes 0.05. In this case, 10,000,000 will 
be used, and this fact reported. 
• Model adequacy will be checked by examination of residuals or other goodness of 
fit tests as needed. Inadequate model fit will prompt us to report alternative models. 
• Multiple imputation will be used in case of missing baseline covariates (if 
applicable). Missing outcome variables will be handled by sensitivity analysis and 
reporting of conditional results. 
 
Supplementary analyses. 
The purpose of the supplementary analyses reported in this section is to assess the 
role of statistical choices and data quality choices in shaping the result. 
 
mortality Fisher exact test 
 
Secondary analyses of mortality. 
All secondary analyses will be sharply distinguished from the primary prespecified 
analysis and will be identified as such. Secondary analyses include outcome variables or 
planned subsets which contribute either additional insight or address different scientific 
questions than the primary analysis. 


 
 
7 
 
A pre-specified secondary analysis will pool infants randomized in the neonate and 
individual studies using a binomial regression with complementary log-log link, permuted 
within study.  A pooled sample size of 54414 infants would have 80% power to detect a 
12% reduction in mortality. 
age Binomial regression with complementary log-log link, using age at treatment (in days) 
 
Prespecified subgroup analyses. 
We will estimate differences separately by the following subgroups, defined by baseline 
characteristics, for both mortality and anthropometry endpoints: 
• age Age at enrollment, by week 
• sex Male versus female 
• season of enrollment Rainy versus dry 
• region Centre, Boucle du Mouhoun, Cascade, Centre Ouest, Haut-Bassins 
• urbanicity Urban versus peri-urban versus rural 
• birthweight  Low birthweight (<2500 g) versus normal birthweight (>= 2500 g) 
• Underweight (WAZ < –2), stunted (LAZ < –2), wasted (WLZ < –2) 
 
Additional secondary outcomes. 
Note that participants outside of WHO Child Growth Standards for WAZ (-6 to +5 SD), 
HAZ (-6 to +6 SD), or WHZ (-5 to +5 SD) will be excluded from all anthropometric 
analyses. 
infantile hypertrophic pyloric stenosis Since the importance of avoiding type II errors 
in safety studies is well known [SL12], we propose to report the one-sided 90% 
confidence interval of the estimated relative risk for IHPS, and tabulate the 
occurrence of IHPS by age at treatment and gender. 


 
 
8 
 
adverse events The total number of individuals reporting an adverse event, the number 
reporting each adverse event; composite serious adverse events. Adverse events 
reported on any of the 7, 14, and 21-day timepoints will be reported in aggregate. 
clinic visit Negative binomial regression on clinic visit counts, for the following reasons given 
for visiting the clinic: Malaria, Pneumonia, Diarrhea, Fever. 
 
hospitalization We will report the two-sided 95% confidence interval of the estimated 
relative risk and tabulate occurrence by age at treatment and gender. 
hospitalization and/or deaths Similarly, we will report the two-sided 95% confidence 
interval of the estimated relative risk and tabulate occurrence by age at treatment 
and gender. 
weight comparison of grams per kilogram per day (growth velocity) between treatment 
and placebo arms, permuting at the level of randomization. 
height comparison of millimeters per day (growth velocity) between treatment and 
placebo arms, permuting at the level of randomization. 
weight, height, MUAC, WHZ, WAZ, HAZ ANCOVA comparing outcomes between 
treatment and placebo arms using baseline as a covariate. 
neonatal mortality Binomial regression with complementary log-log link and firth 
correction used to ensure numerical stability. The analysis will be two-sided, with a 
type I error rate (alpha) of 0.05, analyzed identically to the primary outcome. 
12-month mortality Binomial regression with complementary log-log link and firth 
correction used to ensure numerical stability. The analysis will be two-sided, with a 
type I error rate (alpha) of 0.05, analyzed identically to the primary outcome. 
cause-specific mortality Leveraging verbal autopsy information, specific causes of 
death will be analyzed as binomial regression with complementary log-log link. It 
will be two-sided, with a type I error rate (alpha) of 0.05.


 
 
9 
 
2.2 Interim Monitoring 
 
Efficacy. Interim analysis will be executed by the trial biostatistician at the central site. 
A single interim analysis conducted at alpha of 0.001 will be conducted. Specifically, the 
interim analysis will be conducted when full data are available for the first third of patients 
(six months after the last individual in the first third of patients) or at the end of the first full 
year, whichever occurs first. 
Futility. An interim analysis of futility is proposed, to be finalized in consultation with 
the Data and Safety Monitoring Board. Specifically, we propose to conduct the same 
regression as in the primary analysis comparing the treatment and control arms using 
only data from the first year. This analysis will be conducted using simulation. We suggest 
consideration of the conditional power to detect a 30% effect. If this drops below 10% at 
the interim analysis, discontinuation of the trial or other changes to the protocol may be 
made in consultation with the Data and Safety Monitoring Board. 
Safety. Pyloric stenosis is a rare, though serious, outcome. Cases will be tabulated, 
and the proportion experiencing this outcome by treatment status will be reported on a 
quarterly basis, along with listing of symptoms, onset date, and surgical outcomes once 
performed. We will cross-tabulate by age and gender. Statistical tests will be reported, 
but the decision to discontinue for safety is not expected to be made based solely on 
statistical considerations, but rather in conjunction with the DSMC. 
We will summarize adverse events overall and separately by study arm. We will report 
adverse events by study arm to the DSMC at interim meetings throughout the trial, but 
primary investigators will be masked to the arm-stratified results until the primary outcome 
analysis is unmasked at the end of the trial. We will summarize adverse events 
(numerator) by children at risk (denominator) and will disaggregate and report separately 
more common events following best practice recommendations [LBM+16]. We will 
estimate the risk difference in adverse events between arms and 95 percent confidence 
interval for the difference, though we acknowledge that the trial is not necessarily powered 
to detect differences in adverse events. 


 
 
10 
 
3 Sample Size Considerations 
 
We use the following formula: 
 
𝑛= 𝑝!(1 −𝑝!) + 𝑝"(1 −𝑝")
(𝑝" −𝑝!)#
*𝑧"$%/# −𝑧"$',
# 
 
 
We assume a mortality probability of 0.035. We also assume a reduction in the probability of 
mortality, this reduction being 20%. Specifically, the mortality in the azithromycin group is 
one minus this probability, multiplied by the placebo mortality probability; an effect size 
expressed by 10% corresponds to multiplying the baseline probability by 1-10% = 1-0.1 = 
0.9. 
mortality probability assumed 0.035 (over the study period of six months). 
 
effect size assumed to be 20%. 
 
loss to follow-up assumed to be 10%. 
 
The proposed sample size per arm is 10856, for a total of 21712 in both arms. 
Sensitivity analysis, 20% effect size 
 
• mortality rate 25 per thousand, loss to follow up 10%: total enrollment 30682. 
 
• mortality rate 35 per thousand, loss to follow up 20%: total enrollment 24426. 
 
 


 
 
11 
 
 
Power for secondary outcomes 
All analyses assume a two-sided alpha of 0.05 and a power of 0.8. 
 
nutritional status The proposed sample size will be sufficient to detect a standardized 
effect size of 0.04 for weight and height. 
 
pyloric stenosis The proposed sample size will be sufficient to detect a relative risk of 
approximately 2.1. 


 
 
12 
 
4 Randomization 
 
The randomization will be conducted using R. The function sample with option 
replace=FALSE will be used to conduct the random shuffling. Note that the choice of the random 
number seed completely determines the randomization. To ensure the integrity of the 
randomization, we will use the procedure we used for MORDOR/Malawi. 


 
 
13 
 
5 Abbreviations 
 
ANCOVA Analysis of Covariance 
DSMC Data and Safety Monitoring Committee 
HAZ Height for Age Z score 
HDSS Health and Demographic Surveillance System 
LAZ Length for Age Z score 
MUAC Mid Upper Arm Circumference 
SAP Statistical Analysis Plan 
WAZ Weight for Age Z score  
WHZ Weight for Height Z score 
WLZ Weight for Length Z score


 
 
14 
 
References 
 
[LBM+16] N. Lineberry, J. A. Berlin, B. Mansi, S. Glasser, M. Berkwits, C. Klem, and et al. 
Recommendations to improve adverse event reporting in clinical trial 
publications: a joint pharmaceutical industry/journal editor perspective. British 
Medical Journal, 355, 2016. 
[SL12] S. Singh and Y. K. Loke. Drug safety assessment in clinical trials: methodological 
challenges and opportunities. Trials, 13, 2012. 


 
 
15 
 
6 Revision History 
 
26 Sep 2018 Sample size formula revision. A pre-specified secondary analysis pooling 
infants randomized in the neonate and individual studies was added. 
28 Feb 2019 Revisions to interim, futility, and safety analyses according to Sep 2018 
DSMC recommendations. Addition of secondary endpoints: cause-specific 
mortality, 12- month mortality, and neonatal mortality. 
26 Mar 2019 Added pre-specifications of follow-up date time window and sensitivity 
analysis for primary outcome. 
19 Apr 2019 Clarification regarding unplanned analyses and use of multiple comparison 
corrections. 
26 Jul 2019 Changed mortality analyses from logistic regression to binomial with 
complementary log-log link. 
14 Oct 2019 Added comment that deaths on enrollment day contribute 0.5 days person 
time. 
28 Oct 2019 Revision to futility analysis and secondary outcomes. 
 
09 Dec 2019 Added adverse event detail to the safety component of interim analysis. 
 
16 Jan 2020 Abbreviated the study name on title page. 
 
07 May 2020 Specified WHO standard range for inclusion for anthropometric (child growth)  
analysis. 
21 July 2020 Added secondary analysis of negative binomial regression on clinic visit 
counts. 
14 April 2021 Added prespecified subgroup analyses and aggregate adverse event 
reporting under additional secondary outcomes. 
14 May 2021 Defined subgroup analyses for birthweight, growth standards and regions. 


 
 
16 
9 June 2021 Updated regions for subgroup analyses, added LAZ and WLZ abbreviation 
definitions 
10 June 2021 Added a breakdown of the reasons for visiting the clinic under additional 
secondary outcomes