BackgroundAntibiotic resistance is rising in important bacterial pathogens. Phage therapy (PT), the use of bacterial viruses infecting the pathogen in a species-specific way, is a potential alternative.MethodT4-like coliphages or a commercial Russian coliphage product or placebo was orally given over 4 days to Bangladeshi children hospitalized with acute bacterial diarrhea. Safety of oral phage was assessed clinically and by functional tests; coliphage and Escherichia coli titers and enteropathogens were determined in stool and quantitative diarrhea parameters (stool output, stool frequency) were measured. Stool microbiota was studied by 16S rRNA gene sequencing; the genomes of four fecal Streptococcus isolates were sequenced.FindingsNo adverse events attributable to oral phage application were observed (primary safety outcome). Fecal coliphage was increased in treated over control children, but the titers did not show substantial intestinal phage replication (secondary microbiology outcome). 60% of the children suffered from a microbiologically proven E. coli diarrhea; the most frequent diagnosis was ETEC infections. Bacterial co-pathogens were also detected. Half of the patients contained phage-susceptible E. coli colonies in the stool. E. coli represented less than 5% of fecal bacteria. Stool ETEC titers showed only a short-lived peak and were otherwise close to the replication threshold determined for T4 phage in vitro. An interim analysis after the enrollment of 120 patients showed no amelioration in quantitative diarrhea parameter by PT over standard care (tertiary clinical outcome). Stool microbiota was characterized by an overgrowth with Streptococcus belonging to the Streptococcus gallolyticus and Streptococcus salivarius species groups, their abundance correlated with quantitative diarrhea outcome, but genome sequencing did not identify virulence genes.InterpretationOral coliphages showed a safe gut transit in children, but failed to achieve intestinal amplification and to improve diarrhea outcome, possibly due to insufficient phage coverage and too low E. coli pathogen titers requiring higher oral phage doses. More knowledge is needed on in vivo phage–bacterium interaction and the role of E. coli in childhood diarrhea for successful PT.FundingThe study was supported by a grant from . The trial was registered with Identifier NCT00937274 at ClinicalTrials.gov.

To examine the contributions of impaired gut microbial community development to childhood undernutrition, we combined metabolomic and proteomic analyses of plasma samples with metagenomic analyses of fecal samples to characterize the biological state of Bangladeshi children with severe acute malnutrition (SAM) as they transitioned, after standard treatment, to moderate acute malnutrition (MAM) with persistent microbiota immaturity. Host and microbial effects of microbiota-directed complementary food (MDCF) prototypes targeting weaning-phase bacterial taxa underrepresented in SAM and MAM microbiota were characterized in gnotobiotic mice and gnotobiotic piglets colonized with age- and growth-discriminatory bacteria. A randomized, double-blind controlled feeding study identified a lead MDCF that changes the abundances of targeted bacteria and increases plasma biomarkers and mediators of growth, bone formation, neurodevelopment, and immune function in children with MAM.

The genomic diversity of 99 T4-like coliphages was investigated by sequencing an equimolar mixture with Illumina technology and screening them against different databases for horizontal gene transfer and undesired genes. A 9-phage cocktail was given to 15 healthy adults from Bangladesh at a dose of 3×10(9) and 3×10(7) plaque-forming units and placebo respectively. Phages were detected in 64% of the stool samples when subjects were treated with higher titer phage, compared to 30% and 28% with lower-titer phage and placebo, respectively. No Escherichia coli was present in initial stool samples, and no amplification of phage was observed. One percent of the administered oral phage was recovered from the feces. No adverse events were observed by self-report, clinical examination, or from laboratory tests for liver, kidney, and hematology function. No impact of oral phage was seen on the fecal microbiota composition with respect to bacterial 16S rRNA from stool.

Phage therapy has a long tradition in Eastern Europe, where preparations are comprised of complex phage cocktails whose compositions have not been described. We investigated the composition of a phage cocktail from the Russian pharmaceutical company Microgen targeting Escherichia coli/Proteus infections. Electron microscopy identified six phage types, with numerically T7-like phages dominating over T4-like phages. A metagenomic approach using taxonomical classification, reference mapping and de novo assembly identified 18 distinct phage types, including 7 genera of Podoviridae, 2 established and 2 proposed genera of Myoviridae, and 2 genera of Siphoviridae. De novo assembly yielded 7 contigs greater than 30 kb, including a 147-kb Myovirus genome and a 42-kb genome of a potentially new phage. Bioinformatic analysis did not reveal undesired genes and a small human volunteer trial did not associate adverse effects with oral phage exposure.

Iron absorption from ferrous fumarate was significantly lower than that from ferrous sulfate in both H. pylori-infected and uninfected Bangladeshi children. Treatment of H. pylori infection improved gastric acid output but did not significantly influence iron absorption. The efficacy of ferrous fumarate in iron fortification programs to prevent iron deficiency in young children should be evaluated.

BACKGROUND Environmental enteric dysfunction (EED) is an enigmatic disorder of the small intestine postulated to play a role in childhood undernutrition, a pressing global health problem. Defining the incidence of EED, its pathophysiology, and its contribution to impaired linear and ponderal growth has been hampered by the difficulty in directly sampling the small intestinal mucosa and microbial community (microbiota). METHODS Slum-dwelling Bangladeshi children aged 18±2 months, with linear growth-faltering (stunting) who failed a nutritional intervention underwent endoscopy to obtain duodenal biopsies and aspirates. Levels of 4077 plasma proteins and 2619 duodenal proteins were quantified in 80 children with histopathologic evidence of EED, and the abundances of bacterial strains in their duodenal microbiota were determined using culture-independent methods. Young germ-free mice, fed a Bangladeshi diet, were colonized with bacterial strains cultured from the duodenal aspirates. RESULTS The absolute abundances of a shared group of 14 bacterial strains recovered from the duodenums of children with EED and not typically classified as enteropathogens were negatively correlated with linear growth (length-for-age Z-score;β=-0.38±0.12(SEM);ρ=-0.49;p=0.003), and positively correlated with duodenal proteins involved in immunoinflammatory responses. Representation of these 14 duodenal taxa was significantly different in fecal microbiota from EED versus healthy children (p<0.001;PERMANOVA). Gnotobiotic mice colonized with cultured EED-donor duodenal strains develop a small intestinal enteropathy. CONCLUSIONS These results provide evidence of a causal relationship between components of the small intestinal microbiota, enteropathy and stunting and offer a rationale for developing therapeutics that target what must no longer remain terra incognita-the small intestinal microbiota. ClinicalTrials.gov identifier: NCT02812615

Treatment with antirotavirus immunoglobulin of bovine colostral origin is effective in the management of children with acute rotavirus diarrhea.

Antimicrobial susceptibility of 120 Helicobacter pylori isolates to metronidazole, tetracycline, clarithromycin, and amoxicillin was determined, and 77.5, 15, 10, and 6.6% of the isolates, respectively, were resistant. Only rdxA inactivation and both rdxA and frxA inactivation were responsible for metronidazole resistance in 66% (8 of 12) and 33% (4 of 12) of the isolates, respectively. Eradication ofHelicobacter pylori infection by treatment with two antimicrobial agents (clarithromycin and amoxicillin or metronidazole) and a proton pump inhibitor is recommended by various consensus groups (10,16,20). Antimicrobial resistance in H. pylori is a growing problem as it is the most important factor in determining treatment outcome. The prevalence of antimicrobial resistance varies with geographical regions (3, 25). Metronidazole resistance in H. pylori has been shown to be due to mutation in rdxA; mutation in frxA has also been shown to be associated with metronidazole resistance (11,12,23). In Bangladesh, the prevalences of H. pylori infection among infants, children, and adults are 61, 84, and 92%, respectively (1, 21, 22); however, information on antimicrobial susceptibility to commonly used drugs in H. pylori treatment is lacking. This study was conducted to evaluate (i) the prevalence of primary antibiotic resistance to commonly used antimicrobial agents and (ii) the genetic basis for metronidazole resistance in H. pylori isolates from Bangladesh.Consecutive patients attending the Gastroenterology Department of Dhaka Medical College Hospital for upper gastrointestinal endoscopy were enrolled during 1999 to 2001. Diagnosis of peptic ulcer (PU) and non-ulcer dyspepsia (NUD) or gastritis was based on endoscopic examination of the stomach and duodenum. Biopsy samples were taken from each patient for culture.Bacteria were grown in brain heart infusion agar with 7% sheep blood and incubated at 37°C in 5% O 2 , 10% CO 2 , and 85% N 2 for 3 to 6 days. The MICs of amoxicillin, clarithromycin, metronidazole, and tetracycline for the isolates were determined by the agar dilution method as described elsewhere (18,19). All tests were repeated twice, and H. pylori 26695 was used as a control. ␤-Lactamase production was tested by the chromogenic cephalosporin method (6). The molecular mechanism of susceptibility and resistance to metronidazole was studied in 12 isolates. Metronidazole-susceptible (Mtz s ) isolates were further studied (by inactivation of rdxA alone or rdxA and frxA for conversion into an Mtz r phenotype) by transformation of Mtz s isolates with plasmids pBS-rdxA-cam (rdxA::cat) and pBS-frxA-kan (frxA::kan) as described earlier (11,12).A total of 278 consecutive patients between 15 and 78 years of age were enrolled, and among them, 72.7% (202 patients) were male and 27.3% (76 patients) were female. Among the patients, 162 had PU and 116 had NUD and 62.6% (174 of 278) were culture positive for H. pylori. Among the culturepositive patients, 121 (69.5%) were male and 53 (30.4%) were female and 112 (64.3%) had PU ...