Characterization of Allergenic Fungal Spores from Selected Locations in Lagos and Ibadan, Nigeria

Author: Odebode, Adeyinka John

Supervisors: Adekunle A. A. ; Adeonipekun A. P. and Farombi O. E.

Airborne fungal spores (Air mycoflora) are ubiquitous both indoors and outdoors due to their sizes, predominance and aerodynamic properties which enhance their distribution. Fungal spores are associated with dysfunction of multiple system and organs such as respiratory, nervous, immune, haematological and skin. Hence, this study investigated a two-year comparative survey of airborne fungal spores in parts of Lagos and Ibadan, Nigeria. Sedimentation plate technique using Dichloran Glycerol-18 (DG-18) and Potato Dextrose Agar (PDA) (culture dependent) was employed for fungi collection. Fungi samples were subjected to polymerase chain reaction while the most abundant were screened by quantitative PCR analysis. Histology, total protein content, blood parameters and biochemical profile of oxidation stress markers on fungi inoculated balb/c mice were determined to compare the immune response and effects of different fungi dosage inoculated intranasally. Data obtained were analysed (ANOVA) using SAS (version 9.1). Means were separated using the Duncan Multiple Range Test at p≤0.05. The results revealed species diversity in all the sampling locations. A total of 39 fungal species and strains were isolated from all locations sampled with Aspergillus and Penicillium identified as the most abundant and frequent fungal species in all the surveyed environments while Absidia, Curvularia and Mucor had very low values of spore count in all locations. It was observed that spore count in the dry season was significantly less than that of the wet season, with DG- 18 agar having higher collection (53.8%) than Potato dextrose agar (46.2%). Lagos state also gave more pooled abundant fungi collection than Ibadan. Molecular characterization result of the isolated fungi revealed that no DNA sequence data was 100% homologous with those in the Gene bank. The mean relative gene expression values ranged from 18.95 – 31.28 for Actin, 17.38 – 26.77 for β tubulin, 19.74 – 30.63 for P. oxalicum and 30.22 – 37.56 for P. citrinum allergenic genes. All genes were significantly correlated to the Bestkeeper index (p< 0.001). Histopathology result for all inoculated organisms on mice lung appear to be similar although with varying degrees of severity. Almost all of them had intra-lesional unstained fungal hyphae. Pathologies include thickening of alveolar sepatae, which causes impairment in vascular exchange and/or respiratory movements. Hyperplasia of the bronchiolar epithelium was observed which was caused by the fungal hyphae. The contribution of principal component analysis (PCA) showed variations in the Eigen proportion of fungi inoculum at different concentrations on extent of infection recorded on the lungs of balb/c albino mice inoculated with different fungi treatments. The first principal component analysis accounted for the highest variation with the highest Eigen value for A. flavus at proportion of 38.13%. The fifth component showed positive and more relatedness for control treatment and A. penicilloides. P. citrinum, P. chrysogenum, A. flavus and A. penicilloides produced significant (p<0.05) effect on Glutathione, Superoxide dismutase, Nitric oxide, Hydrogen peroxide and Protein. Fungal treatment is positive and highly significantly (p<0.01) correlated with MPO and Hydrogen peroxide with r=0.66 and 0.38. Blood parameters showed that eosinophils, basophils, monocytes, neutrophils and leucocytes are significantly different from one another for each inoculated organism. Statistical analysis show there was negative but significant relationship between relative humidity and temperature and also a negative relationship exist between temperature and fungi count. Positive and significant correlation occur between asthma and wheezing cough situations.