The chronic rhinosinusitis microbiome

The body of research described within this PhD thesis investigates the presence and role of microbial communities or the microbiome in chronic rhinosinusitis (CRS). After a rigorous review of the literature as it relates to the aetiopathogenesis and microbiology of this disease it is clear that much confusion exists regarding the importance of host and environmental factors which predispose towards the disease state. In particular, significant deficiencies exist in how we conceptualize the role of bacteria and fungi in this ecological niche because our understanding of these microbes has been shaped by detection techniques that lack the sensitivity to accurately detect the microbiome constituents. To address this, we set out to accurately define the microbiome in CRS patients and healthy controls as a basis for further ecological studies. Using this knowledge we then attempted to manipulate the microbiome in a murine model of sinusitis so that the probiotic potential of certain bacteria could be determined. The first study presented in this thesis (Chapter 2) is concerned with the bacteriology of CRS patients as determined through traditional culture results. Here the results from over 500 CRS patients were utilized and several important findings were identified. The important pathogen S. aureus was determined to be the most prevalent species in our cohort, followed by P. aeruginosa, Haemophilus spp. and S. pneumonia. The rates of isolation for S. aureus were also found to be significantly higher in patients undergoing revision surgery, which indicated an important relationship between this organism and recalcitrance. This study formed an important basis for the other microbe detection studies described in the following chapters, which utilized sensitive molecular techniques. Chapter 3 sees investigation of the bacterial component of CRS microbiome using a 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing technique. The large number of species detected in both controls and CRS patients highlighted the sensitivity of this technique for detecting bacteria. Unlike the previous study, other metrics of presence were also characterized such as richness, diversity and relative abundance. The temporal changes occurring in the CRS cohort were examined post-operatively and correlations were made with quality of life. S. aureus was shown to be prevalent in both CRS patients and controls, however its relative abundance in CRS patients was significantly higher than in the control group. The species Acinetobacter johnsonii was also of interest. Here it was found to be significantly more abundant in the control group, which may be of functional importance for the healthy sinonasal microbiome. The study reported in Chapter 4 describes the equally important, but until this time neglected, fungal component of the microbiome. In this instance 18S rDNA fungal tag-encoded FLX amplicon pyrosequencing was performed and fungi were found to be ubiquitous to all patients. The striking finding of this study was the sheer diversity of fungi present across the cohort. Several genera including Malassezia were described for the first time in the sinonasal cavity. The presence of Malassezia was of great interest due to its previously described association with atopic disease. In CRS Malassezia may also play an important role as a disease modifier. Finally, after successfully defining the microbiome our next aim was to manipulate it (Chapter 5). Here we investigated the probiotic potential of S. epidermidis against S. aureus in a murine model of sinusitis. Intranasal inoculations of phosphate buffered saline, S. epidermidis, S, aureus or a combination of the two were administered to 20 mice. PAS-positive goblet cell counts of the mouse nasal mucosa revealed that mice receiving a combination of S. epidermidis and S. aureus had significantly lower goblet cell counts than those receiving S. aureus alone. This finding was of great importance as it confirmed the probiotic potential of S. epidermidis by demonstrating its ability to augment the pathogenicity of S. aureus in the nasal cavity. The implications of this finding for topical intranasal probiotics in CRS are far reaching and should not be underestimated. Most importantly it highlights the viability of ecologically based treatment paradigms in the management of CRS.Thesis (Ph.D.) (Research by Publication) -- University of Adelaide, School of Medicine, 2014