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 these lethal sequelae characteristic of this disease. We report on two published primary studies and a third collaborative work: Our first report features our ongoing evaluation of the immunomodulatory capacity of probiotic Lactobacillus species. We showed previously that wild-type mice primed via intranasal inoculation with live or heat-inactivated Lactobacillus species were fully (100%) protected against the lethal sequelae of infection with the virulent pathogen, pneumonia virus of mice (PVM), a response that is associated with diminished expression of proinflammatory cytokines and diminished virus recovery. We extended these findings with a demonstration that 40% of the mice primed with live Lactobacillus survived when PVM challenge was delayed for as long as 5 months. This robust and sustained resistance to PVM infection resulting from prior interaction with an otherwise unrelated microbe is a profound example of heterologous immunity;we seek to understand the nature and unique features of this response. We found that intranasal inoculation with L. reuteri elicited rapid, transient neutrophil recruitment in association with proinflammatory mediators (CXCL1, CCL3, CCL2, CXCL10, TNF-alpha and IL-17A) but not Th1 cytokines;indeed, the classic Th1 cytokine, interferon-gamma, does not contribute to survival promoted by Lactobacillus-priming. Live L. reuteri detected in lung tissue underwent rapid clearance, and was undetectable at 24h after inoculation. In contrast, L. reuteri peptidoglycan (PGN) and L. reuteri genomic DNA (gDNA) were detected at 24 and 48h after inoculation, respectively. In contrast to live bacteria, intranasal inoculation with isolated L. reuteri gDNA elicited no neutrophil recruitment, had minimal impact on virus recovery and virus-associated production of CCL3, and provided no protection against the negative sequelae of virus infection. Isolated PGN elicited neutrophil recruitment and proinflammatory cytokines but did not promote sustained survival in response to subsequent PVM infection. Overall, further evaluation of the responses leading to Lactobacillus-mediated heterologous immunity may provide insight into novel antiviral preventive modalities. In our second report, we continued with our ongoing evaluation of novel pneumovirus pathogens isolated in the wild by our collaborators at Cornell University College of Veterinary Medicine. The original novel isolate, PnV-Ane4, replicated in and could be re-isolated in infectious state from mouse lung but elicited minimal mortality compared to pneumonia virus of mice (PVM) strain J3666. We previously assessed phylogeny and physiologic responses to 10 new PnV isolates. The G/glycoprotein sequences of all PnVs include elongated amino-termini when compared to the characterized PVMs, and suggest division into groups A and B. While we observed significant differences in cytokine production and neutrophil recruitment to the lungs of BALB/c mice in response to survival doses (50 TCID50 units) of representative group A (114378-10-29-KY-F) and group B (7968-11-OK) PnVs, we observed no evidence for positive selection (dN>dS) among the PnV/PnV, PVM/PnV or PVM/PVM G/glycoprotein or F/fusion protein sequence pairs. These novel variants, while fulfilling Koch's postulates in mice, remain distinct, and provide interesting avenues for exploring responses to unique virus determinants. The final report is a contribution to a collaborative work with the laboratory of Dr. Joseph Domachowske, SUNY Upstate Medical University, Syracuse, NY. This is a clinical study that focuses on the contribution of virus-induced inflammation to the pathogenesis of respiratory syncytial virus (RSV) disease.
The aim of the study was to identify specific mediators that are associated with more severe forms of illness in young children. Children less than 5 yrs of age who were seen in emergency department of Golisano Childrens'Hospital for respiratory symptoms from September 1998 to May 2008 were eligible for enrollment. Nasopharyngeal (NP) wash samples were collected from all eligible patients, and clinical data were recorded. Individuals were included in this study if NP wash samples were positive for RSV only. Patients enrolled in the study were stratified by disease severity, defined as mild (not hospitalized), moderate (hospitalized), or severe (requiring ICU stay). Concentrations of individual inflammatory biomarkers in NP wash fluids were determined using the Luminex human 30-plex assay. 851 patients met study criteria, which included 268 (31.5 percent) with mild, 503 (59.1 percent) with moderate, and 80 (9.4 percent) with severe illness. As expected, illness severity was directly associated with young age, prematurity, heart or lung disease, infection with RSV group A, and elevated concentrations of interleukin (IL)-2R, IL-6, CXCL8, tumor necrosis factor (TNF)-α, interferon (IFN)-α, CCL3, CCL4, and CCL2. In addition, the study identified several novel and mechanistically important inflammatory biomarkers of severe RSV disease, including IL-1β, IL1-RA, IL-7, epidermal growth factor (EGF), and hepatocyte growth factor (HGF).
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