Inflammation and Therapy for Respiratory Virus Infection

National Institute Of Allergy And Infectious Diseases

Investigators

Linked publications & trials

Abstract

Ongoing research focuses on the exploration of pathologic inflammatory responses to acute respiratory virus infection and the use of this information to develop creative strategies to circumvent the lethal sequelae characteristic of this disease. We report three original publications associated with this report. In our first original manuscript, we continue our study of immunobiotic Lactobacillus plantarum (Lp) and its administration to the respiratory mucosa as a means to promote cross-protection against lethal pneumovirus infection. Here, we examine Lp-mediated cross-protection in immunodeficient Rag1-/- mice which cannot clear virus from lung tissue. Interestingly, Lp was protective in this strain. These results confirm that Lp functions primarily as an immumodulatory agent, and can elicit protection in mice that support ongoing virus replication. However, as the Rag1-/- mice cannot clear virus, they do ultimately succumb to a delayed lethal outcome, and repeated administration of Lp to virus-infected Rag1-/- mice had no impact on the delayed response. By contrast, we found that mice devoid of CD8 T cells were fully capable of clearing virus, and were protected by Lp with no delayed lethal outcome, granulocyte recruitment to the airways, or induction of CCL7. Moreover, administration of Lp to the respiratory mucosa, singly or repeatedly, resulted in no induction of IFN-alpha or IFN-beta in wild-type or Rag1-/- or mice, and IFNR gene-deletion had no impact on Lp-mediated protection. Overall, our findings indicate that Lp administered to the respiratory tract has substantial local impact on lethal virus-induced inflammation, regardless of virus clearance. However, endogenous virus clearance mechanisms are still required to promote sustained protection. Citation: Percopo CM, Ma M, Rosenberg HF (2017) Administration of immunobiotic L. plantarum delays but does not prevent lethal respiratory virus infection in lymphocyte-deficient Rag1-/- mice. J. Leukoc. Biol. In press. In our second original manuscript, we examined a prominent assay for detecting bacterial peptidoglycan, and published an important report documenting a critical limitation of this technology. Notably, the silkworm larvae plasma (SLP) assay has been developed as a means to detect bacterial peptidoglycan as a surrogate for live bacteria. In our report, we present results indicating that generation of melanin by this assay is not fully reliable as a surrogate marker for bacterial counts. Notably, we found that melanin levels, the end product of this assay, were not related to absolute bacterial counts. We found that melanin was related local inflammatory responses in lung tissue (probably secondary to protease activation of the SLP cascade), and was directly dependent on signaling via pattern recognition receptors TLR2 and NOD2. This is particularly important given recent interest in this commerically-available assay for use in clinical settings, including primary diagnosis and detection of post-operative infections. Citation: Ma M, Rice TA, Percopo CM, Rosenberg HF (2017) Silkworm Larvae Plasma SLP) assay for detection of bacteria: false positives secondary to inflammation in vivo. J. Microbiol. Methods. 132: 9-13. In our third original manuscript, we generated recombinant green-fluorescent Lactobacillus for our ongoing studies of this agent and its immunomodulatory mechanisms. Using the pTRKH3p15A-ldhGFP plasmid with described by Lizier and colleagues, we used an 8% glycine / glycerol medium to create a stable transfectant cultures from our identified immunobiotic strain of L. reuteri. As shown in this manuscript, we have used this reagent to explore eosinophil-mediated uptake of fluorescently-labeled bacteria, notably (but not limited to) a laboratory strain of E. coli, and the Gram-positive probiotic, Lactobacillus reuteri L275. There are numerous further applications of this assay. Among them, this assay can be extended to include fluorescence-activated cell sorting in order to separate populations and ultimately to distinguish unique gene expression profiles characteristic of the eosinophils that have taken up bacteria (ie., that are GFP-positive) compared to those eosinophils that remain GFP-negative. In this context, it will be also intriguing to explore the impact of specific gene-deletions, notably those related to specific recognition of bacterial patterns and pathogens. Citation: Kraemer LS, Brenner TA, Krumholz JO, Rosenberg HF (2017) . A Flow-Cytometric Method to Evaluate Eosinophil-Mediated Uptake of Probiotic Lactobacillus reuteri. J. Microbiol. Methods 137: 19-24.