Altered gut microbiota in infants is associated with respiratory syncytial virus disease severity.

BACKGROUND: Respiratory syncytial virus (RSV) is the number one cause of lower respiratory tract infections in infants. There are still no vaccines or specific antiviral therapies against RSV, mainly due to the inadequate understanding of RSV pathogenesis. Recent data suggest a role for gut microbiota community structure in determining RSV disease severity. Our objective was to determine the gut microbial profile associated with severe RSV patients, which could be used to help identify at-risk patients and develop therapeutically protective microbial assemblages that may stimulate immuno-protection. RESULTS: We enrolled 95 infants from Le Bonheur during the 2014 to 2016 RSV season. Of these, 37 were well-babies and 58 were hospitalized with RSV. Of the RSV infected babies, 53 remained in the pediatric ward (moderate) and 5 were moved to the pediatric intensive care unit at a later date (severe). Stool samples were collected within 72 h of admission; and the composition of gut microbiota was evaluated via 16S sequencing of fecal DNA. There was a significant enrichment in S24_7, Clostridiales, Odoribacteraceae, Lactobacillaceae, and Actinomyces in RSV (moderate and severe) vs. controls. Patients with severe RSV disease had slightly lower alpha diversity (richness and evenness of the bacterial community) of the gut microbiota compared to patients with moderate RSV and healthy controls. Beta diversity (overall microbial composition) was significantly different between all RSV patients (moderate and severe) compared to controls and had significant microbial composition separating all three groups (control, moderate RSV, and severe RSV). CONCLUSIONS: Collectively, these data demonstrate that a unique gut microbial profile is associated with RSV disease and with severe RSV disease with admission to the pediatric intensive care unit. More mechanistic experiments are needed to determine whether the differences observed in gut microbiota are the cause or consequences of severe RSV disease.

Data on changes to mucosal inflammation and the intestinal microbiota following dietary micronutrients in genetically susceptible hosts.

These data support the findings that dietary micronutrients influence the inflammatory responses and intestinal microbial community structure and function in a model of pouchitis-like small bowel inflammation reported in "Dietary Antioxidant Micronutrients Alter Mucosal Inflammatory Risk in a Murine Model of Genetic and Microbial Susceptibility" (Pierre et al., 2018) [1]. Briefly, wild-type and IL-10 deficient mice underwent surgical placement of small intestinal self-filling loops (SFL) and were subsequently fed purified control diet (CONT) or control diet supplemented with 4 micronutrients (AOX), retinoic acid, Vitamin C, Vitamin E, and selenium, for 14 days. These data include changes in host markers, such as body weight, mucosal levels of myeloperoxidase and syndecan-1, and luminal IgA and IgG levels. These data also include changes in the microbial compartment, including 16S community structure in the self-filling loop, conventionalized germ-free mice, and microbial substrate preference performed through anaerobic bacterial culturing of SLF CONT and AOX microbiota.

Altered neuronal density and neurotransmitter expression in the ganglionated region of Ednrb null mice: implications for Hirschsprung's disease.

BACKGROUND: Hirschsprung's disease (HSCR) is a congenital condition in which enteric ganglia, formed from neural crest cells (NCC), are absent from the terminal bowel. Dysmotility and constipation are common features of HSCR that persist following surgical intervention. This persistence suggests that the portion of the colon that remains postoperatively is not able to support normal bowel function. To elucidate the defects that underlie this condition, we utilized a murine model of HSCR. METHODS: Mice with NCC-specific deletion of Ednrb were used to measure the neuronal density and neurotransmitter expression in ganglia. KEY RESULTS: At the site located proximal to the aganglionic region of P21 Ednrb null mice, the neuronal density is significantly decreased and the expression of neurotransmitters is altered compared with het animals. The ganglia in this colonic region are smaller and more isolated while the size of neuronal cell bodies is increased. The percentage of neurons expressing neuronal nNOS and VIP is significantly increased in Ednrb nulls. Conversely, the percentage of choline acetyltransferase (ChAT) expressing neurons is decreased, while Substance P is unchanged between the two genotypes. These changes are limited to the colon and are not detected in the ileum. CONCLUSIONS & INFERENCES: We demonstrate changes in neuronal density and alterations in the balance of expression of neurotransmitters in the colon proximal to the aganglionic region in Ednrb null mice. The reduced neuronal density and complementary changes in nNOS and ChAT expression may account for the dysmotility seen in HSCR.

Molecular dynamics, crystallography and mutagenesis studies on the substrate gating mechanism of prolyl oligopeptidase.

Altered prolyl oligopeptidase (PREP) activity is found in many common neurological and other genetic disorders, and in some cases PREP inhibition may be a promising treatment. The active site of PREP resides in an internal cavity; in addition to the direct interaction between active site and substrate or inhibitor, the pathway to reach the active site (the gating mechanism) must be understood for more rational inhibitor design and understanding PREP function. The gating mechanism of PREP has been investigated through molecular dynamics (MD) simulation combined with crystallographic and mutagenesis studies. The MD results indicate the inter-domain loop structure, comprised of 3 loops at residues, 189-209 (loop A), 577-608 (loop B), and 636-646 (loop C) (porcine PREP numbering), are important components of the gating mechanism. The results from enzyme kinetics of PREP variants also support this hypothesis: When loop A is (1) locked to loop B through a disulphide bridge, all enzyme activity is halted, (2) nicked, enzyme activity is increased, and (3) removed, enzyme activity is only reduced. Limited proteolysis study also supports the hypothesis of a loop A driven gating mechanism. The MD results show a stable network of H-bonds that hold the two protein domains together. Crystallographic study indicates that a set of known PREP inhibitors inhabit a common binding conformation, and this H-bond network is not significantly altered. Thus the domain separation, seen to occur in lower taxa, is not involved in the gating mechanism for mammalian PREP. In two of the MD simulations we observed a conformational change that involved the breaking of the H-bond network holding loops A and B together. We also found that this network was more stable when the active site was occupied, thus decreasing the likelihood of this transition.

Molecular dynamics study of prolyl oligopeptidase with inhibitor in binding cavity.

We used the crystal structure of prolyl oligopeptidase (POP) with bound Z-pro-prolinal (ZPP) inhibitor (Protein Data Bank (PDB) structure 1QFS) to perform an intensive molecular dynamics study of the POP-ZPP complex. We performed 100 ns of simulation with the hemiacetal bond, through which the ZPP is bound to the POP, removed in order to better investigate the binding cavity environment. From basic analysis, measuring the radius of gyration, root mean square deviation, solvent accessible surface area and definition of the secondary structure of protein, we determined that the protein structure is highly stable and maintains its structure over the entire simulation time. This demonstrates that such long time simulations can be performed without the protein structure losing stability. We found that water bridges and hydrogen bonds play a negligible role in binding the ZPP thus indicating the importance of the hemiacetal bond. The two domains of the protein are bound by a set of approximately 12 hydrogen bonds, specific to the particular POP protein.

A comparison of gut evacuation models for larval mackerel (Scomber scombrus) using serial photography.

A novel technique is described, using serial photography of the gut contents of transparent living larval fishes, to generate individual gut evacuation time series. This technique was applied to Atlantic mackerel Scomber scombrus larvae to compare three widely used models of gut evacuation: linear, exponential and square-root. Regression r(2) for the exponential model exceeded those for the linear and square root models in 20 of 21 time series, strongly supporting the exponential model. At the initial gut fullness for each time series, total gut evacuation rates calculated with the exponential model averaged 2.2 and 1.3 times greater than those calculated with the linear and square-root models, respectively, and would produce correspondingly higher estimates of feeding rates for field-collected larvae with similar levels of gut fullness. The results highlight the importance of choosing the appropriate evacuation model in feeding studies, particularly those intended to examine short-term changes in larval fish feeding rates, a contributing factor to the highly variable yearly recruitment of many marine fish species.

Molecular dynamics simulations of the enzyme catechol-O-methyltransferase: methodological issues.

Results from extensive 70 ns all-atom molecular dynamics simulations of catechol-O-methyltransferase (COMT) enzyme are reported. The simulations were performed with explicit TIP3P water and Mg2+ ions. Four different crystal structures of COMT, with and without different ligands, were used. These simulations are among the most extensive of their kind and as such served as a stability test for such simulations. On the methodological side we found that the initial energy minimization procedure may be a crucial step: particular hydrogen bonds may break, and this can initiate an irreversible loss of protein structure that becomes observable in longer time scales of the order of tens of nanoseconds. This has important implications for both molecular dynamics and quantum mechanics-molecular mechanics simulations.