Copy Number Variation of the Beta Defensin Gene Cluster on Chromosome 8p Influences the Bacterial Microbiota within the Nasopharynx of Otitis-Prone Children

As there is increasing evidence that aberrant defensin expression is related to susceptibility for infectious disease and inflammatory disorders, we sought to determine if copy number of the beta-defensin gene cluster located on chromosome 8p23.1 (DEFB107, 106, 105, 104, 103, DEFB4 and SPAG11), that shows copy number variation as a block, was associated with susceptibility to otitis media (OM). The gene DEFB103 within this complex encodes human beta defensin-3 (hBD-3), an antimicrobial peptide (AP) expressed by epithelial cells that line the mammalian airway, important for defense of mucosal surfaces and previously shown to have bactericidal activity in vitro against multiple human pathogens, including the three that predominate in OM. To this end, we conducted a retrospective case-control study of 113 OM prone children and 267 controls aged five to sixty months. We identified the copy number of the above defined beta-defensin gene cluster (DEFB-CN) in each study subject by paralogue ratio assays. The mean DEFB-CN was indistinguishable between subjects classified as OM prone based on a recent history of multiple episodes of OM and control subjects who had no history of OM (4.4±0.96 versus 4.4±1.08, respectively: Odds Ratio [OR]: 1.16 (95% CI: 0.61, 2.20). Despite a lack of direct association, we observed a statistically significant correlation between DEFB-CN and nasopharyngeal bacterial colonization patterns. Collectively, our findings suggested that susceptibility to OM might be mediated by genetic variation among individuals, wherein a DEFB-CN less than 4 exerts a marked influence on the microbiota of the nasopharynx, specifically with regard to colonization by the three predominant bacterial pathogens of OM.     

Nucleotide-binding oligomerization domain-containing protein 2 regulates suppressor of cytokine signaling 3 expression in Burkholderia pseudomallei-infected mouse macrophage cell line RAW 264.7

Burkholderia pseudomallei, a causative agent of melioidosis, is a facultative intracellular Gram-negative bacterium that can survive and multiply inside the macrophages. Toll-like receptors are one class of pattern recognition receptors (PRRs) that have been documented to play significant role in B. pseudomallei infection. In the present study, we investigated a potential role of nucleotide-binding oligomerization domain-containing protein 1 and 2 (NOD1 and NOD2), cytoplasmic pattern recognition receptors, in B. pseudomallei-infected mouse macrophage cell line RAW 264.7. Both live and heat-killed B. pseudomallei were able to up-regulate NOD1 and NOD2 expression in a time-dependent manner. Marked reduction of a negative regulator, suppressor of cytokine signaling 3 (SOCS3), expression was observed only in B. pseudomallei-infected NOD2-depleted macrophages and not in NOD1-depleted macrophages. The decrease in SOCS3 expression also led to an increase in IFN-γ responsiveness as judged by an enhanced STAT-1 phosphorylation on tyrosine 701 in the B. pseudomallei-infected macrophages. Together, these results suggested that, in addition to using other PRRs to evade macrophage defense, B. pseudomallei may also use NOD2 to regulate a negative regulator like SOCS3.     

Routine Habitat Change: A Source of Unrecognized Transient Alteration of Intestinal Microbiota in Laboratory Mice

The mammalian intestine harbors a vast, complex and dynamic microbial population, which has profound effects on host nutrition, intestinal function and immune response, as well as influence on physiology outside of the alimentary tract. Imbalance in the composition of the dense colonizing bacterial population can increase susceptibility to various acute and chronic diseases. Valuable insights on the association of the microbiota with disease critically depend on investigation of mouse models. Like in humans, the microbial community in the mouse intestine is relatively stable and resilient, yet can be influenced by environmental factors. An often-overlooked variable in research is basic animal husbandry, which can potentially alter mouse physiology and experimental outcomes. This study examined the effects of common husbandry practices, including food and bedding alterations, as well as facility and cage changes, on the gut microbiota over a short time course of five days using three culture-independent techniques, quantitative PCR, terminal restriction fragment length polymorphism (TRFLP) and next generation sequencing (NGS). This study detected a substantial transient alteration in microbiota after the common practice of a short cross-campus facility transfer, but found no comparable alterations in microbiota within 5 days of switches in common laboratory food or bedding, or following an isolated cage change in mice acclimated to their housing facility. Our results highlight the importance of an acclimation period following even simple transfer of mice between campus facilities, and highlights that occult changes in microbiota should be considered when imposing husbandry variables on laboratory animals.