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Infectious keratitis is a leading cause of corneal blindness worldwide with little information known about causative etiologies in Malawi, Africa. This area is resource-limited with ophthalmologist and microbiology services. The Department of Ophthalmology at the Kamuzu College of Health Sciences in Blantyre, Malawi, is a participating site of an international corneal ulcer consortium, capriCORN (Comprehensive Analysis of Pathogens, Resistomes, and Inflammatory-markers in the CORNea). In this study, 50 patients with corneal ulcers were swabbed for pathogen identification using RNA-sequencing. Corneal trauma was reported in 41% and 19% of the patients worked in agriculture. A pathogen was identified in 58% of the cases. Fungal pathogens predominated, followed by viruses and bacteria. Aspergillus, Fusarium, HSV-1, and Gardnerella were the most common pathogens detected. 50% of patients reported treatment with an antibiotic before presentation. Pathogens unusual for infectious keratitis, such as Subramaniula asteroids, Aureobasidium pullulans, and Gardnerella vaginalis, were also detected.

Mass antibiotic distribution to preschool children resulted in alterations of the gut microbiome months after distribution. This individually randomized, placebo-controlled trial evaluated changes in the gut microbiome and resistome in children aged 8 days to 59 months after one dose of oral azithromycin in Burkina Faso. A total of 450 children were randomized in a 1:1 ratio to either placebo or azithromycin. Rectal samples were collected at baseline, 2 weeks, and 6 months after randomization and subjected to DNA deep sequencing. Gut microbiome diversity and normalized antimicrobial resistance determinants for different antibiotic classes were evaluated. Azithromycin decreased gut bacterial diversity (Shannon P < 0.0001; inverse Simpson P < 0.001) 2 weeks after treatment relative to placebo. Concurrently, the normalized abundance of macrolide resistance genetic determinants was 243-fold higher (95% CI: 76-fold to 776-fold, P < 0.0001). These alterations did not persist at 6 months, suggesting that disruptions were transient. Furthermore, we were unable to detect resistance changes in other antibiotic classes, indicating that co-resistance with a single course of azithromycin when treated at the individual level was unlikely.

Wastewater-based surveillance is increasingly recognized as an important approach to monitoring population-level antimicrobial resistance (AMR). In this exploratory study, we examined the use of metagenomics to evaluate AMR using untreated wastewater samples routinely collected by the Niger national polio surveillance program. Forty-eight stored samples from two seasons each year over 4 years (2016-2019) in three regions were selected for inclusion in this study and processed using unbiased DNA deep sequencing. Normalized number of reads of genetic determinants for different antibiotic classes were compared over time, by season, and by location. Correlations in resistance were examined among classes. Changes in reads per million per year were demonstrated for several classes, including decreases over time in resistance determinants for phenicols (-3.3, 95% CI: -8.7 to -0.1, P = 0.029) and increases over time for aminocoumarins (3.8, 95% CI: 0.0 to 11.4, P = 0.043), fluoroquinolones (6.8, 95% CI: 0.0 to 20.5, P = 0.048), and beta-lactams (0.85, 95% CI: 0.1 to 1.7, P = 0.006). Sulfonamide resistance was higher in the post-rainy season compared with the dry season (5.2-fold change, 95% CI: 3.4 to 7.9, P < 0.001). No differences were detected when comparing other classes by season or by site for any antibiotic class. Positive correlations were identified in genetic determinants of resistance among several antibiotic classes. These results demonstrate the potential utility of leveraging existing wastewater sample collection in this setting for AMR surveillance.

The first manifestation of a viral infection may be conjunctivitis. There are limited data on the etiology of viral conjunctivitis in Vanuatu, a country in the South Pacific Ocean. Patients presenting to one of two Vanuatu health centers with presumed infectious conjunctivitis were eligible if symptom onset was within 14 days of screening. Conjunctival and anterior nasal swabs were obtained and subjected to unbiased RNA deep sequencing (RNA-seq) to identify DNA and RNA viruses. For samples collected from May to November 2021, RNA-seq identified a viral etiology in 12/48 patients. Human adenovirus species were the most common viruses (58%) detected, followed by human herpes viruses (cytomegalovirus, varicella zoster virus, and human herpes 7 virus). Rhinovirus C, Epstein-Barr virus, and bocavirus were also detected. In summary, the etiology for viral conjunctivitis in Vanuatu appears broad. Unbiased testing may be useful for disease surveillance.

Antibiotics are routinely used as part of the management of severe acute malnutrition and are known to reduce gut microbial diversity in non-malnourished children. We evaluated gut microbiomes in children participating in a randomized controlled trial (RCT) of azithromycin versus amoxicillin for severe acute malnutrition. Three hundred one children aged 6 to 59 months with uncomplicated severe acute malnutrition (mid-upper arm circumference < 11.5 cm and/or weight-for-height Z-score < -3 without clinical complications) were enrolled in a 1:1 RCT of single-dose azithromycin versus a 7-day course of amoxicillin (standard of care). Of these, 109 children were randomly selected for microbiome evaluation at baseline and 8 weeks. Rectal swabs were processed with metagenomic DNA sequencing. We compared alpha diversity (inverse Simpsons index) at 8 weeks and evaluated relative abundance of microbial taxa using DESeq2. Of 109 children enrolled in the microbiome study, 95 were followed at 8 weeks. We found no evidence of a difference in alpha diversity between the azithromycin and amoxicillin groups at 8 weeks controlling for baseline diversity (mean difference -0.6, 95% CI -1.8 to 0.6, P = 0.30). Gut microbiomes did not diversify during the study. Differentially abundant genera at the P < 0.01 level included Salmonella spp. and Shigella spp., both of which were overabundant in the azithromycin compared with amoxicillin groups. We found no evidence to support an overall difference in gut microbiome diversity between azithromycin and amoxicillin among children with uncomplicated severe acute malnutrition, but potentially pathogenic bacteria that can cause invasive diarrhea were more common in the azithromycin group. Trial Registration: ClinicalTrials.gov NCT03568643.

A broad-spectrum antibiotic, typically amoxicillin, is included in many country guidelines as part of the management of uncomplicated severe acute malnutrition (SAM) in children without overt clinical symptoms of infection. Alternative antibiotics may be beneficial for children with SAM without increasing selection for beta-lactam resistance. We conducted a 1:1 randomized controlled trial of single dose azithromycin versus a 7-day course of amoxicillin for SAM. Children 6-59 months of age with uncomplicated SAM (mid-upper arm circumference < 11.5 cm and/or weight-for-height Z-score < -3) were enrolled in Boromo District, Burkina Faso, from June through October 2020. Rectal swabs were collected at baseline and 8 weeks after treatment and processed using DNA-Seq. We compared the resistome at the class level in children randomized to azithromycin compared with amoxicillin. We found no evidence of a difference in the distribution of genetic antibiotic resistance determinants to any antibiotic class 8 weeks after treatment. There was no difference in genetic macrolide resistance determinants (65% azithromycin, 65% placebo, odds ratio, OR, 1.00, 95% confidence interval, CI, 0.43-2.34) or beta-lactam resistance determinants (82% azithromycin, 83% amoxicillin, OR 0.94, 95% CI, 0.33-2.68) at 8 weeks. Although presence of genetic antibiotic resistance determinants to macrolides and beta-lactams was common, we found no evidence of a difference in the gut resistome 8 weeks after treatment. If there are earlier effects of antibiotics on selection for genetic antibiotic resistance determinants, the resistome may normalize by 8 weeks.

Zoonotic transmission is likely a pathway for antibiotic resistance. Data from a randomized trial of pediatric antibiotic administration were secondarily evaluated to determine if poultry ownership was significantly associated with the presence of gut genetic antibiotic resistance determinants among 118 children in Burkina Faso. Antimicrobial resistance (AMR) determinants were classified using DNA sequencing. We measured the relationship between genetic resistance determinants and chicken ownership using a logistic regression model adjusted for confounding variables. Children in households reporting poultry ownership had four times the odds of tetracycline resistance determinants in the gut compared with those without household poultry (odds ratio [OR]: 4.08, 95% CI: 1.08-15.44, P = 0.04). There was no statistically significant difference found for other antibiotic classes. Understanding the origins of antibiotic resistance may help spur the development of interventions to combat the global AMR crisis.

Campylobacter has emerged as a potential important cause of childhood morbidity in sub-Saharan Africa. Biannual mass azithromycin distribution has previously been shown to reduce all-cause child mortality in sub-Saharan Africa. We conducted a randomized controlled trial in Burkina Faso in which children were randomized in a 1:1 fashion to a 5-day course of azithromycin or placebo to investigate the effect of oral antibiotics on the gut microbiome. We evaluated the changes in the gut microbiome of preschool children treated with azithromycin using metagenomic DNA sequencing. We found that three Campylobacter species were reduced with azithromycin treatment compared with placebo. These results were consistent with other studies that have shown decreases in Campylobacter species after azithromycin treatment, generating the hypothesis that a decrease in Campylobacter may contribute to observations of reduction in mortality following azithromycin distribution.

Infectious conjunctivitis outbreaks remain a public health burden. This study focuses on the pathogen and antimicrobial resistance (AMR) profiles identified in Niger. Sixty-two patients with acute infectious conjunctivitis who presented to health posts were enrolled from December 2021 to May 2022. Nasal and conjunctival swabs were obtained from each patient. Unbiased RNA deep sequencing (RNA-seq) was used to identify associated pathogens. A pathogen was identified in 39 patients (63%; 95% CI, 50-74). Of those, an RNA virus was detected in 23 patients (59%; 95% CI, 43-73). RNA viruses were diverse and included human coronaviruses (HCoVs): SARS-CoV-2, HCoV-229E, HCoV-HKU1, and HCoV-OC43. A DNA virus was identified in 11 patients (28%; 95% CI, 17-44). Of those, four patients had a coinfection with an RNA virus and two patients had a coinfection with both an RNA virus and a bacterium. DNA viruses were predominantly human herpesvirus (cytomegalovirus, Epstein-Barr virus, human herpesvirus 8) and human adenovirus species B, C, and F. Eighteen patients (46%; 95% CI, 32-61) had a bacteria-associated infection that included Haemophilus influenza, Haemophilus aegyptius, Staphylococcus aureus, Streptococcus pneumoniae, and Moraxella spp. Antimicrobial resistance determinants were detected in either the conjunctiva or nasal samples of 20 patients (32%; 95% CI, 22-45) and were found to be more diverse in the nose (Shannon alpha diversity, 1.12 [95% CI, 1.05-1.26] versus 1.02 [95% CI, 1.00-1.05], P = 0.01). These results suggest the potential utility of leveraging RNA-seq to surveil pathogens and AMR for ocular infections.

Antibiotics are routinely used as part of the management of severe acute malnutrition and are known to reduce gut microbial diversity in non-malnourished children. We evaluated gut microbiomes in children participating in a randomized controlled trial (RCT) of azithromycin versus amoxicillin for severe acute malnutrition. Three hundred one children aged 6 to 59 months with uncomplicated severe acute malnutrition (mid-upper arm circumference < 11.5 cm and/or weight-for-height Z-score < -3 without clinical complications) were enrolled in a 1:1 RCT of single-dose azithromycin versus a 7-day course of amoxicillin (standard of care). Of these, 109 children were randomly selected for microbiome evaluation at baseline and 8 weeks. Rectal swabs were processed with metagenomic DNA sequencing. We compared alpha diversity (inverse Simpson's index) at 8 weeks and evaluated relative abundance of microbial taxa using DESeq2. Of 109 children enrolled in the microbiome study, 95 were followed at 8 weeks. We found no evidence of a difference in alpha diversity between the azithromycin and amoxicillin groups at 8 weeks controlling for baseline diversity (mean difference -0.6, 95% CI -1.8 to 0.6, P = 0.30). Gut microbiomes did not diversify during the study. Differentially abundant genera at the P < 0.01 level included Salmonella spp. and Shigella spp., both of which were overabundant in the azithromycin compared with amoxicillin groups. We found no evidence to support an overall difference in gut microbiome diversity between azithromycin and amoxicillin among children with uncomplicated severe acute malnutrition, but potentially pathogenic bacteria that can cause invasive diarrhea were more common in the azithromycin group. Trial Registration: ClinicalTrials.gov NCT03568643.