Jennifer K. Spinler

Human-Derived Probiotic Lactobacillus reuteri Demonstrate Antimicrobial Activities Targeting Diverse Enteric Bacterial Pathogens

Lactobacillus reuteri is a commensal-derived anaerobic probiotic that resides in the human gastrointestinal tract. L. reuteri converts glycerol into a potent broad-spectrum antimicrobial compound, reuterin, which inhibits the growth of gram-positive and gram-negative bacteria. In this study, we compared four human-derived L. reuteri isolates (ATCC 55730, ATCC PTA 6475, ATCC PTA 4659 and ATCC PTA 5289) in their ability to produce reuterin and to inhibit the growth of different enteric pathogens in vitro. Reuterin was produced by each of the four L. reuteri strains and assessed for biological activity. The minimum inhibitory concentration (MIC) of reuterin derived from each strain was determined for the following enteric pathogens: enterohemorrhagic Escherichia coli, enterotoxigenic E. coli, Salmonella enterica, Shigella sonnei and Vibrio cholerae. We also analyzed the relative abilities of L. reuteri to inhibit enteric pathogens in a pathogen overlay assay. The magnitude of reuterin production did not directly correlate with the relative ability of L. reuteri to suppress the proliferation of enteric pathogens. Additional antimicrobial factors may be produced by L. reuteri, and multiple factors may act synergistically with reuterin to inhibit enteric pathogens.

Mechanisms of probiosis and prebiosis: considerations for enhanced functional foods

The technologies of metagenomics and metabolomics are broadening our knowledge of the roles the human gut microbiota play in health and disease. For many years now, probiotics and prebiotics have been included in foods for their health benefits; however, we have only recently begun to understand their modes of action. This review highlights recent advances in deciphering the mechanisms of probiosis and prebiosis, and describes how this knowledge could be transferred to select for enhancing functional foods targeting different populations. A special focus will be given to the addition of prebiotics and probiotics in functional foods for infants and seniors.

Exploring Metabolic Pathway Reconstruction and Genome-Wide Expression Profiling in Lactobacillus reuteri to Define Functional Probiotic Features

The genomes of four Lactobacillus reuteri strains isolated from human breast milk and the gastrointestinal tract have been recently sequenced as part of the Human Microbiome Project. Preliminary genome comparisons suggested that these strains belong to two different clades, previously shown to differ with respect to antimicrobial production, biofilm formation, and immunomodulation. To explain possible mechanisms of survival in the host and probiosis, we completed a detailed genomic comparison of two breast milk–derived isolates representative of each group: an established probiotic strain (L. reuteri ATCC 55730) and a strain with promising probiotic features (L. reuteri ATCC PTA 6475). Transcriptomes of L. reuteri strains in different growth phases were monitored using strain-specific microarrays, and compared using a pan-metabolic model representing all known metabolic reactions present in these strains. Both strains contained candidate genes involved in the survival and persistence in the gut such as mucus-binding proteins and enzymes scavenging reactive oxygen species. A large operon predicted to encode the synthesis of an exopolysaccharide was identified in strain 55730. Both strains were predicted to produce health-promoting factors, including antimicrobial agents and vitamins (folate, vitamin B12). Additionally, a complete pathway for thiamine biosynthesis was predicted in strain 55730 for the first time in this species. Candidate genes responsible for immunomodulatory properties of each strain were identified by transcriptomic comparisons. The production of bioactive metabolites by human-derived probiotics may be predicted using metabolic modeling and transcriptomics. Such strategies may facilitate selection and optimization of probiotics for health promotion, disease prevention and amelioration.

From Prediction to Function Using Evolutionary Genomics: Human-Specific Ecotypes of Lactobacillus reuteri Have Diverse Probiotic Functions

The vertebrate gut symbiont Lactobacillus reuteri has diversified into separate clades reflecting host origin. Strains show evidence of host adaptation, but how host–microbe coevolution influences microbial-derived effects on hosts is poorly understood. Emphasizing human-derived strains of L. reuteri, we combined comparative genomic analyses with functional assays to examine variations in host interaction among genetically distinct ecotypes. Within clade II or VI, the genomes of human-derived L. reuteri strains are highly conserved in gene content and at the nucleotide level. Nevertheless, they share only 70–90% of total gene content, indicating differences in functional capacity. Human-associated lineages are distinguished by genes related to bacteriophages, vitamin biosynthesis, antimicrobial production, and immunomodulation. Differential production of reuterin, histamine, and folate by 23 clade II and VI strains was demonstrated. These strains also differed with respect to their ability to modulate human cytokine production (tumor necrosis factor, monocyte chemoattractant protein-1, interleukin [IL]-1β, IL-5, IL-7, IL-12, and IL-13) by myeloid cells. Microarray analysis of representative clade II and clade VI strains revealed global regulation of genes within the reuterin, vitamin B12, folate, and arginine catabolism gene clusters by the AraC family transcriptional regulator, PocR. Thus, human-derived L. reuteri clade II and VI strains are genetically distinct and their differences affect their functional repertoires and probiotic features. These findings highlight the biological impact of microbe:host coevolution and illustrate the functional significance of subspecies differences in the human microbiome. Consideration of host origin and functional differences at the subspecies level may have major impacts on probiotic strain selection and considerations of microbial ecology in mammalian species.

If you text them, they will come: using the HIV infant tracking system to improve early infant diagnosis quality and retention in Kenya.

Objective:The objective of this study is to evaluate the impact of the HIV Infant Tracking System (HITSystem) for quality improvement of early infant diagnosis (EID) of HIV services. Design and Setting:This observational pilot study compared 12 months of historical preintervention EID outcomes at one urban and one peri-urban government hospital in Kenya to 12 months of intervention data to assess retention and time throughout the EID cascade of care. Participants:Mother–infant pairs enrolled in EID at participating hospitals before (n = 320) and during (n = 523) the HITSystem pilot were eligible to participate. Intervention:The HITSystem utilizes Internet-based coordination of the multistep PCR cycle, automated alerts to trigger prompt action from providers and laboratory technicians, and text messaging to notify mothers when results are ready or additional action is needed. Main outcome measures:The main outcome measures were retention throughout EID services, meeting time-sensitive targets and improving results turn-around time, and increasing early antiretroviral therapy (ART) initiation among HIV-infected infants. Results:The HITSystem was associated with an increase in the proportion of HIV-exposed infants retained in EID care at 9 months postnatal (45.1–93.0% urban; 43.2–94.1% peri-urban), a decrease in turn-around times between sample collection, PCR results and notification of mothers in both settings, and a significant increase in the proportion of HIV-infected infants started on antiretroviral therapy at each hospital(14 vs. 100% urban; 64 vs. 100% peri-urban). Conclusion:The HITSystem maximizes the use of easily accessible technology to improve the quality and efficiency of EID services in resource-limited settings.

Lactobacillus saerimneri and Lactobacillus ruminis: novel human‐derived probiotic strains with immunomodulatory activities

Human-derived lactobacilli were isolated from fecal samples of healthy volunteers. Forty-six isolates from different volunteers were selected and investigated for their immunomodulatory properties. Conditioned medium from each isolate was assessed for its effect on tumor necrosis factor (TNF) production in lipopolysaccharide-activated THP-1 monocytes. Of 46 Lactobacillus isolates, 12 significantly inhibited TNF production in varying magnitude. Lactobacillus strain TH58 displayed the most potent TNF-inhibitory activity (70% inhibition). In contrast, Lactobacillus strain TH14 exhibited immunostimulatory property by activating TNF production in THP-1 monocytes. Lactobacillus TH14 induced nuclear factor-kappaB (NF-kappaB) activation in the absence of lipopolysaccharide stimulation, whereas Lactobacillus TH58 had no effect on NF-kappaB signaling, irrespective of lipopolysaccharide stimulation. Strain TH58 was identified as Lactobacillus saerimneri and strain TH14 as Lactobacillus ruminis by sequence analysis of their 16S rRNA genes. This is the first report of a human isolate of L. saerimneri with TNF-inhibitory activity and L. ruminis, an indigenous species to humans, with TNF stimulatory activity. Our data suggest the potential use of these two strains as immunoprobiotic candidates.

Functional identification in Lactobacillus reuteri of a PocR-like transcription factor regulating glycerol utilization and vitamin B12 synthesis

BackgroundLactobacillus reuteri harbors the genes responsible for glycerol utilization and vitamin B12 synthesis within a genetic island phylogenetically related to gamma-Proteobacteria. Within this island, resides a gene (lreu_1750) that based on its genomic context has been suggested to encode the regulatory protein PocR and presumably control the expression of the neighboring loci. However, this functional assignment is not fully supported by sequence homology, and hitherto, completely lacks experimental confirmation.ResultsIn this contribution, we have overexpressed and inactivated the gene encoding the putative PocR in L. reuteri. The comparison of these strains provided metabolic and transcriptional evidence that this regulatory protein controls the expression of the operons encoding glycerol utilization and vitamin B12 synthesis.ConclusionsWe provide clear experimental evidence for assigning Lreu_1750 as PocR in Lactobacillus reuteri. Our genome-wide transcriptional analysis further identifies the loci contained in the PocR regulon. The findings reported here could be used to improve the production-yield of vitamin B12, 1,3-propanediol and reuterin, all industrially relevant compounds.

Lactobacillus reuteri-Specific Immunoregulatory Gene rsiR Modulates Histamine Production and Immunomodulation by Lactobacillus reuteri

Human microbiome-derived strains of Lactobacillus reuteri potently suppress proinflammatory cytokines like human tumor necrosis factor (TNF) by converting the amino acid l-histidine to the biogenic amine histamine. Histamine suppresses mitogen-activated protein (MAP) kinase activation and cytokine production by signaling via histamine receptor type 2 (H2) on myeloid cells. Investigations of the gene expression profiles of immunomodulatory L. reuteri ATCC PTA 6475 highlighted numerous genes that were highly expressed during the stationary phase of growth, when TNF suppression is most potent. One such gene was found to be a regulator of genes involved in histidine-histamine metabolism by this probiotic species. During the course of these studies, this gene was renamed the Lactobacillus reuteri-specific immunoregulatory (rsiR) gene. The rsiR gene is essential for human TNF suppression by L. reuteri and expression of the histidine decarboxylase (hdc) gene cluster on the L. reuteri chromosome. Inactivation of rsiR resulted in diminished TNF suppression in vitro and reduced anti-inflammatory effects in vivo in a trinitrobenzene sulfonic acid (TNBS)-induced mouse model of acute colitis. A L. reuteri strain lacking an intact rsiR gene was unable to suppress colitis and resulted in greater concentrations of serum amyloid A (SAA) in the bloodstream of affected animals. The PhdcAB promoter region targeted by rsiR was defined by reporter gene experiments. These studies support the presence of a regulatory gene, rsiR, which modulates the expression of a gene cluster known to mediate immunoregulation by probiotics at the transcriptional level. These findings may point the way toward new strategies for controlling gene expression in probiotics by dietary interventions or microbiome manipulation.

Task shifting an inpatient triage, assessment and treatment programme improves the quality of care for hospitalised Malawian children

We aimed to improve paediatric inpatient surveillance at a busy referral hospital in Malawi with two new programmes: (i) the provision of vital sign equipment and implementation of an inpatient triage programme (ITAT) that includes a simplified paediatric severity‐of‐illness score, and (ii) task shifting ITAT to a new cadre of healthcare workers called ‘vital sign assistants’ (VSAs).

Anti-inflammatory Properties of Gastric-derived Lactobacillus plantarum XB7 in the Context of Helicobacter pylori Infection

Helicobacter pylori colonization of the gastric epithelium induces interleukin‐8 (IL‐8) production and inflammation leading to host cell damage. We searched for gastric‐derived Lactobacillus with the ability to suppress H. pylori‐induced inflammation.