Transitions in lung microbiota landscape associate with distinct patterns of pneumonia progression.

The precise microbial determinants driving clinical outcomes in severe pneumonia are unknown. Competing ecological forces produce dynamic microbiota states in health; infection and treatment effects on microbiota state must be defined to improve pneumonia therapy. Here, we leverage our unique clinical setting, which includes systematic and serial bronchoscopic sampling in patients with suspected pneumonia, to determine lung microbial ecosystem dynamics throughout pneumonia therapy. We combine 16S rRNA gene amplicon, metagenomic, and transcriptomic sequencing with bacterial load quantification to reveal clinically-relevant pneumonia progression drivers. Microbiota states are predictive of pneumonia category and exhibit differential stability and pneumonia therapy response. Disruptive forces, like aspiration, associate with cohesive changes in gene expression and microbial community structure. In summary, we show that host and microbiota landscapes change in unison with clinical phenotypes and that microbiota state dynamics reflect pneumonia progression. We suggest that distinct pathways of lung microbial community succession mediate pneumonia progression.

Evolution of the Umbilical Cord Blood Proteome Across Gestational Development.

Neonatal health is dependent on early risk stratification, diagnosis, and timely management of potentially devastating conditions, particularly in the setting of prematurity. Many of these conditions are poorly predicted in real-time by clinical data and current diagnostics. Umbilical cord blood may represent a novel source of molecular signatures that provides a window into the state of the fetus at birth. In this study, we comprehensively characterized the cord blood proteome of infants born between 24 to 42 weeks using untargeted mass spectrometry and functional enrichment analysis. We determined that the cord blood proteome at birth varies significantly across gestational development. Proteins that function in structural development and growth (e.g., extracellular matrix organization, lipid particle remodeling, and blood vessel development) are more abundant earlier in gestation. In later gestations, proteins with increased abundance are in immune response and inflammatory pathways, including complements and calcium-binding proteins. Furthermore, these data contribute to the knowledge of the physiologic state of neonates across gestational age, which is crucial to understand as we strive to best support postnatal development in preterm infants, determine mechanisms of pathology causing adverse health outcomes, and develop cord blood biomarkers to help tailor our diagnosis and therapeutics for critical neonatal conditions.

The effects of antibiotic exposures on the gut resistome during hematopoietic cell transplantation in children.

Antibiotic resistance is a global threat driven primarily by antibiotic use. We evaluated the effects of antibiotic exposures on the gut microbiomes and resistomes of children at high risk of colonization by antibiotic-resistant bacteria. We performed shotgun metagenomic sequencing of 691 serially collected fecal samples from 80 children (<18 years) undergoing hematopoietic cell transplantation. We evaluated the effects of aerobic (cefepime, vancomycin, fluoroquinolones, aminoglycosides, macrolides, and trimethoprim-sulfamethoxazole) and anaerobic (piperacillin-tazobactam, carbapenems, metronidazole, and clindamycin) antibiotic exposures on the diversity and composition of the gut microbiome and resistome. We identified 372 unique antibiotic resistance genes (ARGs); the most frequent ARGs identified encode resistance to tetracyclines (n = 88), beta-lactams (n = 84), and fluoroquinolones (n = 79). Both aerobic and anaerobic antibiotic exposures were associated with a decrease in the number of bacterial species (aerobic, ß = 0.71, 95% CI: 0.64, 0.79; anaerobic, ß = 0.66, 95% CI: 0.53, 0.82) and the number of unique ARGs (aerobic, ß = 0.81, 95% CI: 0.74, 0.90; anaerobic, ß = 0.73, 95% CI: 0.61, 0.88) within the gut metagenome. However, only antibiotic regimens that included anaerobic activity were associated with an increase in acquisition of new ARGs (anaerobic, ß = 1.50; 95% CI: 1.12, 2.01) and an increase in the relative abundance of ARGs in the gut resistome (anaerobic, ß = 1.62; 95% CI: 1.15, 2.27). Specific antibiotic exposures were associated with distinct changes in the number and abundance of ARGs for individual antibiotic classes. Our findings detail the impact of antibiotics on the gut microbiome and resistome and demonstrate that anaerobic antibiotics are particularly likely to promote acquisition and expansion of antibiotic-resistant bacteria.

Gut microbiome is associated with asthma and race in children with food allergy.

BACKGROUND: The composition of the gut microbiome has been associated with development of atopic conditions such as food allergy (FA) and asthma. African American or Black children with FA have higher rate of asthma compared to their White counterparts. OBJECTIVE: We sought to investigate whether the diversity and relative abundance (RA) of gut microbiota is different between children with FA from different racial backgrounds living in the same cities. Furthermore, we aimed to understand whether the difference in the gut microbiota is associated with asthma in children with FA. METHODS: We analyzed and compared the stool microbiome of a cohort of Black and White children with FA by shotgun genomic sequencing. RESULTS: A total of 152 children with IgE-mediated FA enrolled onto FORWARD (Food Allergy Outcomes Related to White and African American Racial Differences); 30 Black and 122 White children were included. The RA of several bacteria was associated with race and asthma. Most notably the RA of Bacteroides thetaiotaomicron, Chlamydia thrachomatis, Parabacteroides goldsteinii, and Bacteroides eggerthii were significantly higher, while the RA of Bifidobacterium sp CAG:754, Parabacterium johnsonii, Bacteroides intestinalis, and Bifidobacterium breve were significantly lower in stool samples of Black children compared to White children. Asthma was associated with lower RA of B breve, Bifidobacterium catenulatum, Prevotella copri, Veilloella sp CAG:933, and Bacteroides plebius, and higher RA of 3 Bacteroides species. CONCLUSIONS: The observed variations in the gut microbiota of Black and White children such as differences in the Bacteroides and Bifidobacterium species along with their association to history of asthma in our cohort is indicative of their potential role in the higher rate of asthma observed among Black children with FA.

Diversity of plant DNA in stool is linked to dietary quality, age, and household income.

Eating a varied diet is a central tenet of good nutrition. Here, we develop a molecular tool to quantify human dietary plant diversity by applying DNA metabarcoding with the chloroplast trnL-P6 marker to 1,029 fecal samples from 324 participants across two interventional feeding studies and three observational cohorts. The number of plant taxa per sample (plant metabarcoding richness or pMR) correlated with recorded intakes in interventional diets and with indices calculated from a food frequency questionnaire in typical diets (ρ = 0.40 to 0.63). In adolescents unable to collect validated dietary survey data, trnL metabarcoding detected 111 plant taxa, with 86 consumed by more than one individual and four (wheat, chocolate, corn, and potato family) consumed by >70% of individuals. Adolescent pMR was associated with age and household income, replicating prior epidemiologic findings. Overall, trnL metabarcoding promises an objective and accurate measure of the number and types of plants consumed that is applicable to diverse human populations.

Gut microbiota analyses of cutaneous T-cell lymphoma patients undergoing narrowband ultraviolet B therapy reveal alterations associated with disease treatment.

Recent studies have shown a close relationship between cutaneous T-cell lymphoma (CTCL) and its microbiome. CTCL disease progression is associated with gut dysbiosis and alterations in bacterial taxa parallel those observed in immunologically similar atopic dermatitis. Moreover, the microbial profile of lesional skin may predict response to narrowband ultraviolet B (nbUVB), a common skin-directed therapy. However, the relationship between the gut microbiome, an immunologically vital niche, and nbUVB remains unexplored in CTCL. Herein, we performed 16S rRNA sequencing and PICRUSt2 predictive metagenomics on DNA extracted from stool swabs of 13 CTCL patients treated with nbUVB, 8 non-treated patients, and 13 healthy controls. Disease response was assessed with modified Severity Weighted Assessment Tool (mSWAT); of nbUVB-treated patients, 6 improved (decreased mSWAT), 2 remained stable, and 5 worsened (increased mSWAT). Protective commensal bacteria including Lactobacillaceae and Erysipelatoclostridiaceae were significantly less abundant in CTCL patients compared to controls. With treatment, the CTCL gut microbiome exhibited decreased phylogenetic diversity and lower relative abundance of pro-inflammatory Sutterellaceae. Sutterellaceae was also significantly more abundant in patients who worsened, and Eggerthellaceae and Erysipelotrichaceae trended higher in patients who improved. Finally, PICRUSt2 functional predictions based on shifts in abundance of bacterial sequences repeatedly identified alterations in inositol degradation, which plays a key role in host immunomodulation, including inositol phospholipid signaling relevant to T-cell survival and proliferation. Our results bolster the paradigm of gut dysbiosis in CTCL and its functional implications in disease pathogenesis, and further delineate bacterial taxa associated with nbUVB response and with nbUVB treatment itself.

Narrowband ultraviolet B response in cutaneous T-cell lymphoma is characterized by increased bacterial diversity and reduced Staphylococcus aureus and Staphylococcus lugdunensis.

Skin microbiota have been linked to disease activity in cutaneous T-cell lymphoma (CTCL). As the skin microbiome has been shown to change after exposure to narrowband ultraviolet B (nbUVB) phototherapy, a common treatment modality used for CTCL, we performed a longitudinal analysis of the skin microbiome in CTCL patients treated with nbUVB. 16S V4 rRNA gene amplicon sequencing for genus-level taxonomic resolution, tuf2 amplicon next generation sequencing for staphylococcal speciation, and bioinformatics were performed on DNA extracted from skin swabs taken from lesional and non-lesional skin of 25 CTCL patients receiving nbUVB and 15 CTCL patients not receiving nbUVB from the same geographical region. Disease responsiveness to nbUVB was determined using the modified Severity Weighted Assessment Tool: 14 (56%) patients responded to nbUVB while 11 (44%) patients had progressive disease. Microbial α-diversity increased in nbUVB-responders after phototherapy. The relative abundance of Staphylococcus, Corynebacterium, Acinetobacter, Streptococcus, and Anaerococcus differentiated nbUVB responders and non-responders after treatment (q<0.05). Microbial signatures of nbUVB-treated patients demonstrated significant post-exposure depletion of S. aureus (q=0.024) and S. lugdunensis (q=0.004) relative abundances. Before nbUVB, responder lesional skin harboured higher levels of S. capitis (q=0.028) and S. warneri (q=0.026) than non-responder lesional skin. S. capitis relative abundance increased in the lesional skin of responders (q=0.05) after phototherapy; a similar upward trend was observed in non-responders (q=0.09). Post-treatment skin of responders exhibited significantly reduced S. aureus (q=0.008) and significantly increased S. hominis (q=0.006), S. pettenkoferi (q=0.021), and S. warneri (q=0.029) relative abundances compared to that of no-nbUVB patients. Staphylococcus species abundance was more similar between non-responders and no-nbUVB patients than between responders and no-nbUVB patients. In sum, the skin microbiome of CTCL patients who respond to nbUVB is different from that of non-responders and untreated patients, and is characterized by shifts in S. aureus and S. lugdunensis. Non-responsiveness to phototherapy may reflect more aggressive disease at baseline.

Nasal Dysbiosis in Cutaneous T-Cell Lymphoma Is Characterized by Shifts in Relative Abundances of Non-Staphylococcus Bacteria.

The nasal microbiome of patients with cutaneous T-cell lymphoma (CTCL) remains unexplored despite growing evidence connecting nasal bacteria to skin health and disease. Nasal swabs from 45 patients with CTCL (40 with mycosis fungoides, 5 with Sézary syndrome) and 20 healthy controls from the same geographical region (Chicago Metropolitan Area, Chicago, IL) were analyzed using sequencing of 16S ribosomal RNA and tuf2 gene amplicons. Nasal α-diversity did not differ between mycosis fungoides/Sézary syndrome and healthy controls (Shannon index, genus level, P = 0.201), but distinct microbial communities were identified at the class (R2 = 0.104, P = 0.023) and order (R2 = 0.0904, P = 0.038) levels. Increased relative abundance of the genera Catenococcus, Vibrio, Roseomonas, Acinetobacter, and unclassified Clostridiales was associated with increased skin disease burden (P < 0.005, q < 0.05). Performed to accurately resolve nasal Staphylococcus at the species level, tuf2 gene amplicon sequencing revealed no significant differences between mycosis fungoides/Sézary syndrome and healthy controls. Although S. aureus has been shown to worsen CTCL through its toxins, no increase in the relative abundance of this taxon was observed in nasal samples. Despite the lack of differences in Staphylococcus, the CTCL nasal microbiome was characterized by shifts in numerous other bacterial taxa. These data add to our understanding of the greater CTCL microbiome and provide context for comprehending nasal-skin and host‒tumor‒microbial relationships.

A specialized multi-disciplinary care program for children with sepsis and multiple organ dysfunction-associated immune dysregulation.

The complex physiology and medical requirements of children with sepsis and multiple organ dysfunction syndrome (MODS) challenge traditional care coordination models. While the involvement of multiple clinical subspecialty services is often necessary to support different care processes and individual organ system dysfunctions, it can also delay the diagnostic process, monitoring, and treatment. The logistics of coordinating with many specialty providers for critically ill patients are challenging and time consuming, and often can result in fragmented communication. To address these and other related issues, we developed a new multi-disciplinary consult service focused on streamlining diagnostics, management, and communication for patients with sepsis and MODS-associated immune dysregulation. The service, called the Program in Inflammation, Immunity, and the Microbiome (PrIIMe), is now a hospital-wide clinical consult service at our institution caring for a broad group of patients with immune dysregulation, particularly focusing on patients with sepsis and MODS. In this paper, we summarize the development, structure, and function of the program, as well as the initial impact. This information may be helpful to clinicians and healthcare leaders who are developing multi-disciplinary consult services for children with complex care needs, especially those with sepsis and MODS-associated immune dysregulation. IMPACT: The care of children with sepsis and multiple organ dysfunction-associated immune dysregulation requires rapid and flexible involvement of multiple clinical subspecialists that is difficult to achieve without fragmented care and delayed decision making. In this narrative review we describe the development, structure, and function of a multi-disciplinary consult service at a children's hospital dedicated to helping coordinate management and provide continuity of care for patients with sepsis and multiple organ dysfunction-associated immune dysregulation. This information may be helpful to clinicians and healthcare leaders who are developing multi-disciplinary consult services for children with complex care needs, especially those with sepsis and MODS-associated immune dysregulation.

Non-diphtheriae Corynebacterium species are associated with decreased risk of pneumococcal colonization during infancy.

Streptococcus pneumoniae (pneumococcus) is a leading cause of severe infections among children and adults. Interactions between commensal microbes in the upper respiratory tract and S. pneumoniae are poorly described. In this study, we sought to identify interspecies interactions that modify the risk of S. pneumoniae colonization during infancy and to describe development of the upper respiratory microbiome during infancy in a sub-Saharan African setting. We collected nasopharyngeal swabs monthly (0-6 months of age) or bimonthly (6-12 months of age) from 179 mother-infant dyads in Botswana. We used 16S ribosomal RNA gene sequencing to characterize the nasopharyngeal microbiome and identified S. pneumoniae colonization using a species-specific PCR assay. We detect S. pneumoniae colonization in 144 (80%) infants at a median age of 71 days and identify a strong negative association between the relative abundance of the bacterial genera Corynebacterium within the infant nasopharyngeal microbiome and the risk of S. pneumoniae colonization. Using in vitro cultivation experiments, we demonstrate growth inhibition of S. pneumoniae by secreted factors from strains of several Corynebacterium species isolated from these infants. Finally, we demonstrate that antibiotic exposures and the winter season are associated with a decline in the relative abundance of Corynebacterium within the nasopharyngeal microbiome, while breastfeeding is associated with an increase in the Corynebacterium relative abundance. Our findings provide novel insights into the interspecies interactions that contribute to colonization resistance to S. pneumoniae and suggest that the nasopharyngeal microbiome may be a previously unrecognized mechanism by which environmental factors influence the risk of pneumococcal infections during childhood. Moreover, this work lays the foundation for future studies seeking to use targeted manipulation of the nasopharyngeal microbiome to prevent infections caused by S. pneumoniae.

Advanced Obesity Treatment Selection among Adolescents in a Pediatric Weight Management Program.

Background: Treatment options for adolescents with obesity are limited. Yet, therapies previously reserved for adults, such as medications and bariatric surgery, are increasingly available to adolescents in tertiary obesity treatment settings. We aimed to identify the factors associated with selecting an advanced obesity treatment (diets, medications, and surgery) beyond lifestyle therapy among adolescents presenting to a tertiary, pediatric weight management program. Methods: We conducted a secondary analysis of adolescents (N = 220) who participated in a longitudinal, observational case-control study within a pediatric weight management program. The exposures were potential individual and clinical factors, including sociodemographic characteristics and comorbidities. The outcome was treatment selection, dichotomized into lifestyle vs. advanced treatment. We modeled associations between these factors and treatment selection using logistic regression, controlling for confounding variables (age, race/ethnicity, sex, and insurance). Results: The study population included a majority of non-Hispanic Black (50.5%) and Hispanic/Latino (19.5%) adolescents, of whom 25.5% selected advanced treatment. Adolescents were more likely to choose an advanced treatment option if they had a greater BMI [odds ratio (OR) 1.09, 95% confidence interval (95% CI) 1.04-1.15], lived further from the clinic (OR 1.03, 95% CI 1.00-1.05), and had an elevated glycohemoglobin level (OR 2.46, 95% CI 1.24-4.92). Conclusions: A significant fraction of adolescents seeking obesity treatment in a specialized care setting chose new and emerging obesity treatments, particularly those at high risk of developing diabetes. These findings can inform patient-clinician obesity treatment discussions in specialty care settings. Clinical Trial Registration number: NCT03139877.

Screening Students and Staff for Asymptomatic Coronavirus Disease 2019 in Chicago Schools.

OBJECTIVES: To assess rates of asymptomatic severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) positivity in K-8 schools with risk mitigation procedures in place, and to evaluate SARS-CoV-2 transmission in school and household contacts of these positive individuals. STUDY DESIGN: In this prospective observational study, screening testing for SARS-CoV-2 was performed by oropharyngeal swabbing and polymerase chain reaction (PCR) analysis in students and staff at K-8 private schools in high-risk Chicago ZIP codes. New coronavirus disease 2019 (COVID-19) diagnoses or symptoms among participants, household contacts, and nonparticipants in each school were queried. RESULTS: Among 11 K-8 private schools across 8 Chicago ZIP codes, 468 participants (346 students, 122 staff members) underwent screening testing. At the first school, 17 participants (36%) tested positive, but epidemiologic investigation suggested against in-school transmission. Only 5 participants in the subsequent 10 schools tested positive for an overall 4.7% positivity rate (1.2% excluding school 1). All but 1 positive test among in-person students had high PCR cycle threshold values, suggesting very low SARS-CoV-2 viral loads. In all schools, no additional students, staff, or household contacts reported new diagnoses or symptoms of COVID-19 during the 2 weeks following screening testing. CONCLUSIONS: We identified infrequent asymptomatic COVID-19 in schools in high-risk Chicago communities and did not identify transmission among school staff, students, or their household contacts. These data suggest that COVID-19 mitigation procedures, including masking and physical distancing, are effective in preventing transmission of COVID-19 in schools. These results may inform future strategies for screening testing in K-8 schools.

Biotransformation of Doxorubicin Promotes Resilience in Simplified Intestinal Microbial Communities.

Chemotherapeutic drugs can cause harmful gastrointestinal side effects, which may be modulated by naturally occurring members of our microbiome. We constructed simplified gut-associated microbial communities to test the hypothesis that bacteria-mediated detoxification of doxorubicin (i.e., a widely used chemotherapeutic) confers protective effects on the human microbiota. Mock communities composed of up to five specific members predicted by genomic analysis to be sensitive to the drug or resistant via biotransformation and/or efflux were grown in vitro over three generational stages to characterize community assembly, response to perturbation (doxorubicin exposure), and resilience. Bacterial growth and drug concentrations were monitored with spectrophotometric assays, and strain relative abundances were evaluated with 16S rRNA gene sequencing. Bacteria with predicted resistance involving biotransformation significantly lowered concentrations of doxorubicin in culture media, permitting growth of drug-sensitive strains in monoculture. Such protective effects were not produced by strains with drug resistance conferred solely by efflux. In the mixed communities, resilience of drug-sensitive members depended on the presence and efficiency of transformers, as well as drug exposure concentration. Fitness of bacteria that were resistant to doxorubicin via efflux, though not transformation, also improved when the transformers were present. Our simplified community uncovered ecological relationships among a dynamic consortium and highlighted drug detoxification by a keystone species. This work may be extended to advance probiotic development that may provide gut-specific protection to patients undergoing cancer treatment. IMPORTANCE While chemotherapy is an essential intervention for treating many forms of cancer, gastrointestinal side effects may precede infections and risks for additional health complications. We developed an in vitro model to characterize key changes in bacterial community dynamics under chemotherapeutic stress and the role of bacterial interactions in drug detoxification to promote microbiota resilience. Our findings have implications for developing bio-based strategies to promote gut health during cancer treatment.

The Pediatric Obesity Microbiome and Metabolism Study (POMMS): Methods, Baseline Data, and Early Insights.

OBJECTIVE: The purpose of this study was to establish a biorepository of clinical, metabolomic, and microbiome samples from adolescents with obesity as they undergo lifestyle modification. METHODS: A total of 223 adolescents aged 10 to 18 years with BMI ≥95th percentile were enrolled, along with 71 healthy weight participants. Clinical data, fasting serum, and fecal samples were collected at repeated intervals over 6 months. Herein, the study design, data collection methods, and interim analysis-including targeted serum metabolite measurements and fecal 16S ribosomal RNA gene amplicon sequencing among adolescents with obesity (n = 27) and healthy weight controls (n = 27)-are presented. RESULTS: Adolescents with obesity have higher serum alanine aminotransferase, C-reactive protein, and glycated hemoglobin, and they have lower high-density lipoprotein cholesterol when compared with healthy weight controls. Metabolomics revealed differences in branched-chain amino acid-related metabolites. Also observed was a differential abundance of specific microbial taxa and lower species diversity among adolescents with obesity when compared with the healthy weight group. CONCLUSIONS: The Pediatric Metabolism and Microbiome Study (POMMS) biorepository is available as a shared resource. Early findings suggest evidence of a metabolic signature of obesity unique to adolescents, along with confirmation of previously reported findings that describe metabolic and microbiome markers of obesity.

Bacterial Signatures of Paediatric Respiratory Disease: An Individual Participant Data Meta-Analysis.

Introduction: The airway microbiota has been linked to specific paediatric respiratory diseases, but studies are often small. It remains unclear whether particular bacteria are associated with a given disease, or if a more general, non-specific microbiota association with disease exists, as suggested for the gut. We investigated overarching patterns of bacterial association with acute and chronic paediatric respiratory disease in an individual participant data (IPD) meta-analysis of 16S rRNA gene sequences from published respiratory microbiota studies. Methods: We obtained raw microbiota data from public repositories or via communication with corresponding authors. Cross-sectional analyses of the paediatric (<18 years) microbiota in acute and chronic respiratory conditions, with >10 case subjects were included. Sequence data were processed using a uniform bioinformatics pipeline, removing a potentially substantial source of variation. Microbiota differences across diagnoses were assessed using alpha- and beta-diversity approaches, machine learning, and biomarker analyses. Results: We ultimately included 20 studies containing individual data from 2624 children. Disease was associated with lower bacterial diversity in nasal and lower airway samples and higher relative abundances of specific nasal taxa including Streptococcus and Haemophilus. Machine learning success in assigning samples to diagnostic groupings varied with anatomical site, with positive predictive value and sensitivity ranging from 43 to 100 and 8 to 99%, respectively. Conclusion: IPD meta-analysis of the respiratory microbiota across multiple diseases allowed identification of a non-specific disease association which cannot be recognised by studying a single disease. Whilst imperfect, machine learning offers promise as a potential additional tool to aid clinical diagnosis.

Microbiology of Bloodstream Infections in Children After Hematopoietic Stem Cell Transplantation: A Single-Center Experience Over Two Decades (1997-2017).

BACKGROUND: Bloodstream infections (BSIs) occur frequently after hematopoietic stem cell transplantation (HSCT). We examined the microbiology of BSI in pediatric HSCT recipients over a 2-decade period at our institution to inform empirical antimicrobial prescribing and infection prevention strategies. METHODS: We conducted a retrospective cohort study of children (<18 years) who underwent HSCT at Duke University between 1997 and 2015. We used recurrent-event gap-time Cox proportional hazards models to determine the hazards of all-cause and cause-specific BSI according to HSCT year. We compared the median time to BSI by causative organism type and evaluated for temporal trends in the prevalence of antibiotic resistance among causative organisms. RESULTS: A total of 865 BSI occurred in 1311 children, including 412 (48%) Gram-positive bacterial, 196 (23%) Gram-negative bacterial, 56 (6%) fungal, 23 (3%) mycobacterial, and 178 (21%) polymicrobial BSI. The hazard of all BSIs did not change substantially over time during the study period, but the hazard of fungal BSIs declined over time during the study period (P = .04). Most fungal BSIs (82%) occurred in the first 100 days after HSCT, whereas mycobacterial BSIs occurred later after HSCT than BSIs caused by other organisms (P < .0001). The prevalence of vancomycin resistance among BSIs caused by Enterococcus faecium increased during the study period (P = .0007). The risk of 2-year mortality in children was increased with BSI (P = .02), Gram-negative bacterial BSI (P = .02), and fungal BSI (P < .0001). CONCLUSIONS: Despite expanded practices for BSI prevention over the past several decades, the incidence of BSI remains high in pediatric HSCT recipients at our institution. Additional strategies are urgently needed to effectively prevent BSIs in this high-risk population.

Optimization of a Noncanonical Anti-infective: Interrogation of the Target Binding Pocket for a Small-Molecule Inhibitor of Escherichia coli Polysaccharide Capsule Expression.

We previously identified a small-molecule inhibitor of capsule biogenesis (designated DU011) and identified its target as MprA, a MarR family transcriptional repressor of multidrug efflux pumps. Unlike other proposed MprA ligands, such as salicylate and 2,4-dinitrophenol (DNP), DU011 does not alter Escherichia coli antibiotic resistance and has significantly enhanced inhibition of capsule expression. We hypothesized that the potency and the unique action of DU011 are due to novel interactions with the MprA binding pocket and the conformation assumed by MprA upon binding DU011 relative to other ligands. To understand the dynamics of MprA-DU011 interaction, we performed hydrogen-deuterium exchange mass spectrometry (HDX-MS); this suggested that four peptide regions undergo conformational changes upon binding DU011. We conducted isothermal calorimetric titration (ITC) to quantitatively characterize MprA binding to DU011 and canonical ligands and observed a distinct two-site binding isotherm associated with the binding reaction of MprA to DU011; however, salicylate and DNP showed a one-site binding isotherm with lower affinity. To elucidate the binding pocket(s) of MprA, we selected single point mutants of MprA that included mutated residues predicted to be within the putative binding pocket (Q51A, F58A, and E65D) as well as on or near the DNA-binding domain (L81A, S83T, and T86A). Our ITC studies suggest that two of the tested MprA mutants had lower affinity for DU011: Q51A and F58A. In addition to elucidating the MprA binding pocket for DU011, we studied the binding of these mutants to salicylate and DNP to reveal the binding pockets of these canonical ligands.

Short-Chain Fatty Acid Production by Gut Microbiota from Children with Obesity Differs According to Prebiotic Choice and Bacterial Community Composition.

Pediatric obesity remains a public health burden and continues to increase in prevalence. The gut microbiota plays a causal role in obesity and is a promising therapeutic target. Specifically, the microbial production of short-chain fatty acids (SCFA) from the fermentation of otherwise indigestible dietary carbohydrates may protect against pediatric obesity and metabolic syndrome. Still, it has not been demonstrated that therapies involving microbiota-targeting carbohydrates, known as prebiotics, will enhance gut bacterial SCFA production in children and adolescents with obesity (age, 10 to 18 years old). Here, we used an in vitro system to examine the SCFA production by fecal microbiota from 17 children with obesity when exposed to five different commercially available over-the-counter (OTC) prebiotic supplements. We found microbiota from all 17 patients actively metabolized most prebiotics. Still, supplements varied in their acidogenic potential. Significant interdonor variation also existed in SCFA production, which 16S rRNA sequencing supported as being associated with differences in the host microbiota composition. Last, we found that neither fecal SCFA concentration, microbiota SCFA production capacity, nor markers of obesity positively correlated with one another. Together, these in vitro findings suggest the hypothesis that OTC prebiotic supplements may be unequal in their ability to stimulate SCFA production in children and adolescents with obesity and that the most acidogenic prebiotic may differ across individuals.IMPORTANCE Pediatric obesity remains a major public health problem in the United States, where 17% of children and adolescents are obese, and rates of pediatric "severe obesity" are increasing. Children and adolescents with obesity face higher health risks, and noninvasive therapies for pediatric obesity often have limited success. The human gut microbiome has been implicated in adult obesity, and microbiota-directed therapies can aid weight loss in adults with obesity. However, less is known about the microbiome in pediatric obesity, and microbiota-directed therapies are understudied in children and adolescents. Our research has two important findings: (i) dietary prebiotics (fiber) result in the microbiota from adolescents with obesity producing more SCFA, and (ii) the effectiveness of each prebiotic is donor dependent. Together, these findings suggest that prebiotic supplements could help children and adolescents with obesity, but that these therapies may not be "one size fits all."

Anaerobic Antibiotics and the Risk of Graft-versus-Host Disease after Allogeneic Hematopoietic Stem Cell Transplantation.

Certain anaerobic bacteria are important for maintenance of gut barrier integrity and immune tolerance and may influence the risk of graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (HSCT). We conducted a single-center retrospective cohort study of allogeneic HSCT recipients to evaluate associations between receipt of antibiotics with an anaerobic spectrum of activity and GVHD outcomes. We identified 1214 children and adults who developed febrile neutropenia between 7 days before and 28 days after HSCT and compared GVHD risk and mortality among patients who received anaerobic antibiotics (piperacillin-tazobactam or carbapenems; n = 491) to patients who received only antibiotics with minimal activity against anaerobes (aztreonam, cefepime, or ceftazidime; n = 723). We performed metagenomic sequencing of serial fecal samples from 36 pediatric patients to compare the effects of specific antibiotics on the gut metagenome. Receipt of anaerobic antibiotics was associated with higher hazards of acute gut/liver GVHD (hazard ratio [HR], 1.26; 95% confidence interval [CI], 1.03 to 1.54) and acute GVHD mortality (HR, 1.63; 95% CI, 1.08 to 2.46), but not chronic GVHD diagnosis (HR, 1.04; 95% CI: .84 to 1.28) or chronic GVHD mortality (HR, .88; 95% CI, .53 to 1.45). Anaerobic antibiotics resulted in decreased gut bacterial diversity, reduced abundances of Bifidobacteriales and Clostridiales, and loss of bacterial genes encoding butyrate biosynthesis enzymes from the gut metagenome. Acute gut/liver GVHD was preceded by a sharp decline in bacterial butyrate biosynthesis genes with antibiotic treatment. Our findings demonstrate that exposure to anaerobic antibiotics is associated with increased risks of acute gut/liver GVHD and acute GVHD mortality after allogeneic HSCT. Use of piperacillin-tazobactam or carbapenems should be reserved for febrile neutropenia cases in which anaerobic or multidrug-resistant infections are suspected.