Characterisation of the Microbial Community in Indoor Environments: a Chemical-Analytical Approach

University dissertation from Department of Laboratory Medicine, Lund University

Abstract: Inhalation of airborne microorganisms and their toxic components and products may induce a wide spectrum of acute and chronic airway manifestations. The study describes important capabilities of chemical marker analysis in defining human exposure to microorganisms in indoor environments. An integrated procedure is presented where gas chromatography - ion trap tandem mass spectrometry is used to determine chemical markers of gram-negative bacterial lipopolysaccharide (3-hydroxy fatty acids with 10-16 carbon atoms), gram-positive bacteria (branched-chain fatty acids with 15 and 17 carbon atoms), bacterial peptidoglycan (muramic acid), and fungal biomass (ergosterol). A hydrolysate of 13C-labelled cyanobacterial cells was used as an internal standard for the first three markers (13C-labelled 3-hydroxytridecanoic acid from labelled Pectinatus cerevisiiphilus was used as an alternative internal standard to improve quantification of the 3-hydroxy fatty acids) and dehydrocholesterol served as an internal standard for ergosterol. These analyses required two 1-5 mg dust samples, one for 3-hydroxy fatty acids, non-hydroxylated fatty acids and muramic acid, and another for ergosterol. The method was used to distinguish between microbial communities in settled and air-borne house dust samples collected from different locations indoors. Both quantitative and qualitative (marker profiles) differences were detected among the studied locations. Air concentrations of 3-hydroxytetradecanoic acid were 4-63 times higher in rooms of smoking students than in parallel rooms of non-smoking students demonstrating that cigarette smoke may be a major source of endotoxin in indoor environments. Settled dust (collected on the floor and from surfaces above the floor) did not show different amounts of lipopolysaccharide with regard to smoking. A school survey demonstrated that the levels of muramic acid were much higher in occupied rooms than in the same rooms when unoccupied. Notably, the levels of 3-hydroxy fatty acids were only slightly higher in occupied than unoccupied rooms suggesting an alteration in bacterial population. Also the particle size distribution in schoolrooms was shown to be quite different in occupied than in unoccupied rooms. This thesis has resulted in the development of a universally applicable method for characterising microbial populations in matrices that are complex chemically. This method forms a basis for epidemiological studies on health consequences upon microbial exposure in indoor environments.