Characterization of Indoor Environment Dust Using Culturable , Chemical, & Molecular Techniques

1. Characterization of Indoor Environment Dust Using Culturable, Chemical, & Molecular Techniques Vinita Katiyar * & Anubha Joshi *INDIRA GANDHI NATIONAL OPEN UNIVERSITY (IGNOU) Regional Centre Karnal 132001 HRYANA Email: vinita.katiyar@gmail.com BACKGROUND Organic dust comprising of viable & non-viable constituents of microorganisms & fractions of plant origin (Millner et al., 1994). Microorganisms (bacteria & fungi), their metabolites & debris considered significant part of the organic dust as causal agents of respiratory disorder & lung dysfunction in occupational (indoor) & non-occupational environment. The measurement of microbial fraction of organic dust in indoor environment samples are of great concern due to their potential negative impact on occupational safety & health. Therefore, it is an important to have an access to adequate methods for enumeration of bacteria & fungi as well as their metabolites present in indoor environment dust (settled & migrated dust). This presentation consists of overview of various techniques of enumeration of bacteria & fungi of environment dust found in various indoor facilities. ANALYTICAL TECHNIQUES These techniques include conventional technique as well as analysis of chemical& biological markers (metabolites) of microbial origins. Limitations: Conventional method is useful for qualitative & quantitative analysis bacteria & fungi of dust. Simultaneously, it has some limitations, since a small fraction (0.1-1%) of the total microflora in an indoor environment dust sample can be detected by this method. Moreover, microscopy does not provide comprehensive information about the microbial characteristics of the collected bioparticles. This method is not fit for the detection of non-culturable microbial community. CONVENTIONAL TECHNIQUE Viable microorganisms are metabolically active (living) organisms with the potential to reproduce. On the basis of ability of reproduction, viable microorganisms may be divided in two subgroups: culturable & nonculturable. Culturablemicroorganisms reproduce under controlled laboratory conditions (In-vitro) using culture technique, which is now known as old & traditional means for detection, enumeration & identification of culturablebacteria & fungi of dust. Technique involves series of steps, i.e., culturing on proper & suitable culture media, colony characteristics, examination of morphological & microscopic features. Non-culturable organisms do not reproduce in the laboratory because of intracellular stress or because the conditions (e.g., culture medium or incubation temperature) are not conducive to growth. Figure 1 & 2 show the quantitative analysis of settled & migrated dust collected from various schools in Delhi (Katiyar 2007). CHEMICAL MARKER ANALYSIS Characterization of indoor organic dust, using standard chemical markers for different groups of microorganisms i.e. markers of bacteria (e.g. endotoxins, LPSpeptidoglycan,muramic acid etc.) & fungal biomarkers (e.g. mycotoxinseargosterol, β-glucans etc.) is a significant & recent approach in this context, since these markers have been found to be associated with respiratory diseases & building-related symptoms (Chao et al 2003, Raoet al 2005). Figure 4:MA content of indoor dust. Quantification of chemical markers of organic dust settled on various indoor surfaces (Delhi & Sweedenbuldings) viz. Lipid polysaccharides (LPS), Muramic acid (Muraminsyra/MA), Ergostecrol, were carried out by in University of Lund, Sweden using Gas Chromatography-Mass Spectroscopy (GC-MS) techniques (Khare & Katiyar 2011) [Figure 3, 4, & 5]. Figure 5: LPS, MA, Ergosterol of Swedish house dust BIOLOGICAL MARKER ANALYSIS In the progression of modern day technology, diagnostic bacteriology, virology, & mycology are rapidly adopting molecular biology techniques in addition to classical identification methods to identify the etiologic agent. Thereby various diagnostic assays utilizing nucleic acid or DNA probes have now been developed for the detection of numerous microorganisms of environmental dusts (Meklin et al 2004, Janke et al (2013). The high specificity, sensitivity & the short period within which the assay can be completed, makes the use of probes advantageous over culture and serology for application in laboratory diagnosis. Polymerase Chain Reaction (PCR) & DNA hybridization techniques nowadays are techniques that are complementary to the conventional detection methods, targeting the DNA of microbes which detect both viable and nonviable (non-reproducible) microbes. On the other hand, the ability to distinguish between microbial strains using various typing procedures based on use of labeled probes like restriction fragment length polymorphism analysis, Random Amplified Polymorphic DNA (RAPD) analysis helps in locating the source of infection. Figure 3: Ergosterol content of indoor dust. References Chao H.J. et al (2003).Environ. Health Perspect., 111, 1242-1248. Janke T. et al (2013). Curr Microbiol. 2013 Mar 10 (Abstruct). Katiyar Vinita (2007). Assessment of biocontaminants from indoor environment Research Project Under Fast Track-Young Scientist Scheme, DST, Govt. of India(SR/L-82/2003) Khare M & Katiyar V. (2011).Article. COOLING INDIA pp 80-83. Mekin T. et al (2004). Environ Monit. 6 (7):615-20. Millner, P. et al (1994). Compost Science & Utilization. 2(4): 6-57. Rao C.Y. et al (2005). Indoor Air, 15 (9) 89-97. Poster Presented in National Conference on "CLIMATE CHANGE : SOCIO-ECONOMIC AND ENVIRONMENTAL ISSUES - PROBLEM AND CHALLENGES" atDepartment of Botany, Meerut College Meerut (UP)on21st & 22nd April,2013.