Antibiotic Resistance and Microbiome in Children Aged 6-59 Months in Nouna, Burkina Faso

This study is designed to better understand the effect of a short course of antibiotics on changes in intestinal and nasopharyngeal microbiome on treated children and untreated household contacts. The investigators hypothesize that a short course of antibiotics will lead to decreased bacterial diversity shortly after completion of the antibiotic course, and higher probability of identification of bacterial resistance genes in rectal and nasopharyngeal samples. The investigators hypothesize that a 5-day course of antibiotics (azithromycin, amoxicillin, or co-trimoxazole) will lead to significantly decreased intestinal and nasopharyngeal bacterial diversity among children aged 6-59 months.

Specific Aim 1. Determine the effect of treatment with antibiotics on microbiome diversity in children aged 6-59 months following a 5-day course of antibiotics.

Specific Aim 1A. Determine the direct effect of a 5-day course of azithromycin, amoxicillin, or co-trimoxazole on intestinal and nasopharyngeal bacterial diversity in children aged 6-59 months compared to no treatment.

Specific Aim 1B. Determine the indirect effect of antibiotic treatment of children in a household on intestinal and nasopharyngeal bacterial diversity in an untreated child aged 6-59 months.

Specific Aim 1C. Assess the association between intestinal bacterial diversity and anthropometry in a population-based sample of children.

• Azithromycin (Drug), also known as Zithromax: Children in this arm will receive Azithromycin once a day.
• Amoxicillin (Drug): Children in this arm will receive Amoxicillin twice a day.
• Cotrimoxazole (Drug): Children in this arm will receive co-trimoxazole once a day.
• Placebo (Drug): Children in this arm will receive Placebo once a day.

Primary outcomes

• Simpson's Index of Diversity (Alpha Diversity) in Intestinal Microbiome [Baseline and Day 9]: The primary outcome of the study was pre-specified as α-diversity (inverse Simpson's) at the genus level, expressed in effective number. Simpson's Alpha Diversity were obtained at Baseline and Post-treatment in this study. The minimum of Simpson's index of diversity is 0, there is no maximum. Higher Simpson's index of diversity means more diverse. There are no subscales.

Secondary outcomes

• Simpson's Index of Diversity (Alpha Diversity) in Microbiome [Day 9]: Direct and indirect effect of antibiotics on alpha diversity from rectal samples
• Weight-for-height Z-score [Day 35]: Nutritional status as determined by weight-for-height Z-score vs. Placebo household Weight-for-height Z-score in each antibiotic group compared with placebo 4 weeks after last antibiotic dose Weight-for-height Z (WHZ) scores were calculated based on the 2006 World Health Organization (WHO) standards. The mean of the 2006 population standards is 0. Lower standard deviations = worse outcomes. A cutoff of < -2 means moderately wasted (WHZ). A cutoff of < -3 means wasted (WHZ).
• Height-for-age Z-score [Day 35]: Nutritional status as determined by height-for-age Z-score Height-for-age Z-score in each antibiotic group compared with placebo 4 weeks after last antibiotic dose Height-for-age Z (HAZ) score were calculated based on the 2006 World Health Organization (WHO) standards. The mean of the 2006 population standards is 0. Lower standard deviations = worse outcomes. A cutoff of < -2 means moderately stunted (HAZ). A cutoff of < -3 means severely stunted (HAZ).
• Weight-for-age Z-score [Day 35]: Nutritional status as determined by weight-for-age Z-score vs. Placebo household Weight-for-age Z-score in each antibiotic group compared with placebo 4 weeks after last antibiotic dose Weight-for-age Z-score (WAZ) scores were calculated based on the 2006 World Health Organization (WHO) standards. The mean of the 2006 population standards is 0. Lower standard deviations = worse outcomes. A cutoff of < -2 means moderately underweight (WAZ). A cutoff of < -3 means severely underweight (WAZ).
• Mid-upper Arm Circumference [Day 35]: Nutritional status as determined by mid-upper arm circumference in each antibiotic group compared with placebo 4 weeks after last antibiotic dose Mid-upper arm circumference (MUAC) in each antibiotic group compared with placebo 4 weeks after last antibiotic dose. MUAC is a measure to assess nutritional status. It is measured on a straight left arm, mid-way between the tip of the shoulder and the tip of the elbow. It identifies acute malnutrition and is commonly used in children 6-59 months of age as well as pregnant women. MUAC less than 115 mm indicates severe wasting or severe acute malnutrition (SAM). MUAC greater than or equal to 115 mm and less than 125 mm indicates moderate wasting or moderate acute malnutrition (MAM).
• Shannon's Index of Diversity (Alpha Diversity) in Intestinal Microbiome [Baseline and Day 9 (Post- Treatment)]: Shannon's Alpha Diversity at Baseline and Post-treatment. combines richness and diversity. Shannon's index of diversity (alpha diversity) measures both the number of species and the inequality between species abundances. A large value is given by the presence of many species with well balanced abundances.
• Shannon's Index of Diversity (Alpha Diversity) in Nasopharyngeal Microbiome [Day 9]: Direct and indirect effects of antibiotics on Shannon's index of bacterial diversity
• L1-norm Distance on Bacterial Reads (Intestinal) [Baseline and Day 9 (Post- Treatment)]: L1-norm distance on bacterial reads (intestinal) - L1 norm is equivalent to Shannon's diversity. Shannon's Alpha Diversity combines richness and diversity. Shannon's index of diversity (alpha diversity) measures both the number of species and the inequality between species abundances. A large value is given by the presence of many species with well balanced abundances.
• L1-norm Distance on Bacterial Reads (Nasopharyngeal) [Day 9]: L1-norm distance on bacterial reads (nasopharyngeal)
• L2-norm Distance on Bacterial Reads (Intestinal) [Baseline and Day 9 (Post- Treatment)]: L2-norm distance on bacterial reads (intestinal) - L2 norm is equivalent to Simpson's diversity. Simpson's Alpha Diversity were obtained at Baseline and Post-treatment in this study. The minimum of Simpson's index of diversity is 0, there is no maximum. Higher Simpson's index of diversity means more diverse. There are no subscales.
• L2-norm Distance on Bacterial Reads (Nasopharyngeal) [Day 9]: L2-norm distance on bacterial reads (nasopharyngeal)
• Number of Participants With Macrolide Resistance Genes [2 years]: Prevalence of macrolide resistance genes measured using DNA-seq from rectal swabs.
• Alpha Diversity in the Intestinal Microbiome [2 years]: Alpha diversity in the intestinal microbiome using DNA-seq from rectal swabs

Sex: All, ages: 6 Months to 59 Months, accepts healthy volunteers

Inclusion Criteria:

* Households will be eligible for inclusion in the study if they have 2 or more children aged 6 months to 59 months currently residing in the household. Children from the household will be eligible if they are 6-59 months of age and are not currently receiving antibiotic treatment

Exclusion Criteria:

* Children who are allergic to any of the study antibiotics will be excluded. Individuals aged under 6 months and 5 years or older will be excluded. Children already receiving antibiotics for an ongoing disease will be excluded.

Participant flow

Outcome results

• Simpson's Index of Diversity (Alpha Diversity) in Intestinal Microbiome (Primary), Mean, Index score: Amoxicillin 10.2 (8.8 to 11.5); Azithromycin 8.8 (7.5 to 10.1); Cotrimoxazole 9.7 (8.2 to 11.2); Placebo 9.6 (8.6 to 10.7)
• Simpson's Index of Diversity (Alpha Diversity) in Microbiome (Secondary)
• Weight-for-height Z-score (Secondary), Mean, z score: Amoxicillin -0.24 +/- 0.92; Azithromycin -0.71 +/- 0.98; Cotrimoxazole -0.27 +/- 0.91; Placebo
• Height-for-age Z-score (Secondary), Mean, z score: Amoxicillin -0.80 +/- 1.19; Azithromycin -0.92 +/- 1.13; Cotrimoxazole -0.69 +/- 1.00; Placebo
• Weight-for-age Z-score (Secondary), Mean, Z score: Amoxicillin -0.60 +/- 0.87; Azithromycin -1.01 +/- 0.95; Cotrimoxazole -0.58 +/- 0.94; Placebo
• Mid-upper Arm Circumference (Secondary), Mean, mm: Amoxicillin 15.01 +/- 0.92; Azithromycin 14.53 +/- 1.11; Cotrimoxazole 15.49 +/- 0.98; Placebo
• Shannon's Index of Diversity (Alpha Diversity) in Intestinal Microbiome (Secondary), Mean, Index score: Amoxicillin 16.6 (14.5 to 18.7); Azithromycin 14.6 (13.0 to 16.2); Cotrimoxazole 15.6 (13.4 to 17.8); Placebo 15.4 (14.1 to 16.7)
• Shannon's Index of Diversity (Alpha Diversity) in Nasopharyngeal Microbiome (Secondary)
• L1-norm Distance on Bacterial Reads (Intestinal) (Secondary), Mean, Index score: Amoxicillin 16.6 (14.5 to 18.7); Azithromycin 14.6 (13.0 to 16.2); Cotrimoxazole 15.6 (13.4 to 17.8); Placebo 15.4 (14.1 to 16.7)
• L1-norm Distance on Bacterial Reads (Nasopharyngeal) (Secondary)
• L2-norm Distance on Bacterial Reads (Intestinal) (Secondary), Mean, Index score: Amoxicillin 10.2 (8.8 to 11.5); Azithromycin 8.8 (7.5 to 10.1); Cotrimoxazole 9.7 (8.2 to 11.2); Placebo 9.6 (8.6 to 10.7)
• L2-norm Distance on Bacterial Reads (Nasopharyngeal) (Secondary)
• Number of Participants With Macrolide Resistance Genes (Secondary), Count of participants, Participants: Azithromycin Households 12; Placebo Households 9
• Alpha Diversity in the Intestinal Microbiome (Secondary)

Adverse events

Time frame: days 1-5

Other adverse events by term

• Results publication: Oldenburg CE, Hinterwirth A, Worden L, Sie A, Dah C, Ouermi L, Coulibaly B, Zhong L, Chen C, Ruder K, Lietman TM, Keenan JD, Doan T. Indirect effect of oral azithromycin on the gut resistome of untreated children: a randomized controlled trial. Int Health. 2021 Feb 24;13(2):130-134. doi: 10.1093/inthealth/ihaa029. PMID 32556194
• Results publication: Dennis EG, Sie A, Ouermi L, Dah C, Tapsoba C, Zabre P, Barnighausen T, O'Brien KS, Lebas E, Keenan JD, Oldenburg CE. Short-term weight gain among preschool children in rural Burkina Faso: a secondary analysis of a randomised controlled trial. BMJ Open. 2019 Jul 29;9(7):e029634. doi: 10.1136/bmjopen-2019-029634. PMID 31362969
• Results publication: Oldenburg CE, Sie A, Coulibaly B, Ouermi L, Dah C, Tapsoba C, Barnighausen T, Lebas E, Arzika AM, Cummings S, Zhong L, Lietman TM, Keenan JD, Doan T. Indirect Effect of Azithromycin Use on the Intestinal Microbiome Diversity of Untreated Children: A Randomized Trial. Open Forum Infect Dis. 2019 Feb 6;6(3):ofz061. doi: 10.1093/ofid/ofz061. eCollection 2019 Mar. PMID 30895203
• Results publication: Oldenburg CE, Sie A, Coulibaly B, Ouermi L, Dah C, Tapsoba C, Barnighausen T, Ray KJ, Zhong L, Cummings S, Lebas E, Lietman TM, Keenan JD, Doan T. Effect of Commonly Used Pediatric Antibiotics on Gut Microbial Diversity in Preschool Children in Burkina Faso: A Randomized Clinical Trial. Open Forum Infect Dis. 2018 Nov 2;5(11):ofy289. doi: 10.1093/ofid/ofy289. eCollection 2018 Nov. PMID 30515431
• Results publication: Sie A, Dah C, Ouermi L, Tapsoba C, Zabre P, Barnighausen T, Lebas E, Arzika AM, Snyder BM, Porco TC, Lietman TM, Keenan JD, Oldenburg CE. Effect of Antibiotics on Short-Term Growth among Children in Burkina Faso: A Randomized Trial. Am J Trop Med Hyg. 2018 Sep;99(3):789-796. doi: 10.4269/ajtmh.18-0342. Epub 2018 Jul 12. PMID 30014828

• Study Protocol (PDF, 2017-05-19)
• Statistical Analysis Plan (PDF, 2017-06-01)

• Thomas M Lietman, MD, Principal investigator, UCSF F.I. Proctor Foundation
• Catherine E Oldenburg, ScD, Study director, UCSF F.I. Proctor Foundation