Isolation and Identification of Fungi from Clinical Samples of Diabetic Patients and Studying the Anti-Fungal Activity of Some Natural Oils on Isolated Fungi
Article Sidebar
Main Article Content
Abstract
Isolation of fungi was performed from February to July, 2019. One hundred clinical specimens were collected from King Abdullah Hospital (KAH) Bisha, Saudi Arabia. Samples were collected from twenty patients of different ages (30 - 70 years old) ten males and ten females. The samples were collected from patients with the two types of diabetics. Specimens included blood, hair, nail, oral swabs and skin. Specimens were inoculated on Sabourauds Dextrose agar containing chloramphenicol. Thirteen fungal species were isolated and identified. The isolated species were: Aspergillus flavus, A. niger, A. terrus, A. nidulans, A. fumigatus, Candida albicans, C. krusei, C. parapsilosis, C. Tropicalis, Curvularia lunata, Fusarium solani, Penicillium marneffei and Saccharomyces cerevisiae. Identification of molds was carried out morphologically and microscopically using available methods and books of identification, while identification of yeasts was carried out using API system. C. albicans recorded the highest isolated number where 31 colonies were isolated from 18 patients, representing relative density of 22.5%. (R. D.: is the number of a certain fungal species divided by the total number of fungi). Other isolated fungal species recorded relative density less than 16 %. The most common isolated fungus Candida albicans was molecularly identified using the 5.8S and flanking ITS regions. The antifungal activity of some natural essential oils (cinnamon, thyme, coconut, almond and clove) was assayed against isolated fungi using disk diffusion method. The used concentration was 5 µl / plate. The MIC values were also determined using different oil concentrations (1, 2.5, 5, 10, 20 and 40 µl / disc).
Received 21/8/2020, Accepted 13/12/2020, Published Online First 21/2/2021
Article Details
This work is licensed under a Creative Commons Attribution 4.0 International License.
How to Cite
References
Hyde KD, Al-Hatmi AMS, Andersen B, Boekhout T, Buzina W, Dawson TL Jr, et al. The world’s ten most feared fungi. Fungal Divers. 2018; 9:161-94.
Rudramurthy SM, Paul RA, Chakrabarti A, Mouton JW, Jacques F, Meis JF. Invasive Aspergillosis by Aspergillus flavus: Epidemiology, Diagnosis, Antifungal Resistance, and Management. J. Fungi. 2019; 5:55-77.
Nami S, Aghebati-Maleki A, Morovati H, Aghebati-Maleki L. Current antifungal drugs and immunotherapeutic approaches as promising strategies to treatment of fungal diseases. Biomed Pharmacother. 2019; 110: 857-68.
Wiederhold NP. Antifungal resistance: current trends and future strategies to combat. Infect Drug Resist. 2017; 10: 249-59.
Khairat SM, Sayed AM, Nabih M, Soliman NS, Hassan YM. Prevalence of Candida blood stream infections among children in tertiary care hospital: detection of species and antifungal susceptibility. Infect Drug Resist. 2019; 12: 2409-16.
Cortés JA, Corrales IF. Invasive candidiasis: epidemiology and risk factors. InFungal Infection 2018 Nov 10. IntechOpen.
Chen LM, Xu YH, Zhou CL, Zhao J, Li CY, Wang R. Overexpression of CDR1 and CDR2 genes plays an important role in fluconazole resistance in Candida albicans with G487T and T916C mutations. J Int Med Res. 2010; 38: 536-45.
Pfaller MA, Diekema DJ, Colombo AL, Kibbler C, Ng KP, Gibbs DL, et al. Candida rugosa, an emerging fungal pathogen with resistance to azoles: geographic and temporal trends from the ARTEMIS DISK Antifungal Surveillance Program. J Clin Microbiol. 2006; 44:3578-82.
Al-Zuhairi AFH. Isolation and identification of pathogenic fungi from diabetic patients in Diyala. Biochem. Cell Arch. 2018; 18(1):959-66.
Santhosh YL, Ramanath KV, Naveen MR. Fungal Infections in Diabetes Mellitus: An Overview. Int J Pharmaceut Sci Review and Research. 2011; 7(2): 221-5.
Hillson R. Fungi and diabetes. Practical Diabetes, 2016; 33(5): 151-2.
Vignesh KB, Amar KG, Swapna M, Joshy ME. Isolation and identification of Candida species from various clinical samples in a tertiary care hospital. Int J Res Med Sci. 2017; 5(8):3520-2.
Pappas PG, Kauffman CA, Andes DR, Clancy CJ, Marr K, Ostrosky-Zeichner LL, et al. Executive Summary: Clinical Practice Guideline for the Management of Candidiasis: 2016 Update by the Infectious Diseases Society of America. Clin Infect Dis. 2016; 62: 409-17.
Whaley SG, Berkow EL, Rybak JR, Nishimoto AT, Katherine S. Barker KS, et al. Azole Antifungal Resistance in Candida albicans and Emerging Non-albicans Candida Species. Front Microbiol. 2016; 7: 2173.
Berkow E, Lockhard S. Fluconazole resistance in Candida species: A current perspective. Infect Drug Resist. 2017; 10:237-45.
Ermis E, Yilmaz A, Boyraz N. Investigation of in vitro and in vivo antifungal activities of different plant essential oils against postharvest apple rot deseases Colletotrichum gleosporioides, Botrytis cinerea and Penicillium expansum. Arch Lebensmittelhyg. 2016; 67(5):122-31.
Nazzaro F, Fratianni F, Coppola R, De Feo V. Essential Oils and Antifungal Activity. Pharmaceuticals. 2017; 10(4):86.
Hoang MTV, Irinyi L, Chen SCA, Sorrell TC, Meyer W. Dual DNA barcoding for the molecular identification of the agents of invasive fungal infections. Front Microbiol. 2019; 10:1647.
Hering D, Borja A, Jones JI, Pont D, Boets P, Bouchez A, et al. Implementation options for DNA-based identification into ecological status assessment under the European Water Framework Directive. Water Res. 2018; 138:182-205.
Wu B, Hussain M, Zhang W, Stadler M, Liu X, Xiang M. Current insights into fungal species diversity and perspective on naming the environmental DNA sequences of fungi. Mycol. 2019; 10(3):127-40.
Letchuman S. Short Introduction of DNA Barcoding. Int J Res. 2018; 5(4): 673-86.
Fajarningsih ND. Internal Transcribed Spacer (ITS) as DNA Barcoding to Identify Fungal Species: a Review. Squalene Bull Mar Fish Post Harvest Biotech. 2016; 11(2): 37-44.
Cheng T, Xu C, Lei L, Li C, Zhang Y, Zhou S, et al. Barcoding the kingdom Plantae: New PCR primers for ITS regions of plants with improved universality and specificity. Mol Ecol Resour. 2016; 16: 138-49.
Maadon SN, Wakid SA, Zainudin II, Rusli LS, Mohdzan MS, Hasan N, et al. Isolation and Identification of Endophytic Fungi from UiTM Reserve Forest, Negeri Sembilan. Sains Malays. 2018; 47(12): 3025-30.
Samson RA, Visagie CM, Houbraken J, Hong SB, Hubka V, Klaassen CHW, et al. Phylogeny, identification and nomenclature of the genus Aspergillus. Studies in mycology. 2014; 78: 141-73.
Larone DH. Medically important fungi: a guide to identification. 4th ed. Washington (D.C.): ASM press, 2002. XVIII, 409 p.
Davey KG, Chant PM, Downer CS, Campbell CK, Warnock DW. Evaluation of the AUXACOLOR system, a new method of clinical yeast identification. J Clin Pathol. 1995; 48:807-9.
Nweze EI, Mukherjee PK, Ghannoum MA. Agar-Based Disk Diffusion Assay for Susceptibility Testing of Dermatophytes. J Clinic Microbiol. 2010; 48(1):3750-2.
Andrews JA. Determination of minimum inhibitory concentrations. Suppl. S1. 2001; 5-16.
Melo SCO, Pungartnik C, Cascardo JCM, Brendel M. Rapid and efficient protocol for DNA extraction and molecular identification of the Basidiomycete Crinipellis perniciosa. Genet Mol Res. 2006; 5(4):851-5.
Korabecna M. The Variability in the Fungal Ribosomal DNA (ITS1, ITS2, and 5.8 S rRNA Gene): Its Biological Meaning and Application. Inin Medical Mycology. In Communicating Current Research and Educational Topics and Trends in Applied Microbiology. Mendez-Vilas (Ed.)2007. 783-787.
Duncan, DB. Multiple ranges and multiple. F-test Biometrics. 1955; 11(1):1- 42.
Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, et al. Gapped BLAST and PSI-BLAST: a new generation of protein database search program. Nucleic Acids Res. 1997; 25(17): 3389 - 3402.
Marr KA, Carter RA, Crippa F, Wald A, Corey L. Epidemiology and outcome of mould infections in hematopoietic stem cell transplant recipients. Clin Infect Dis. 2002; 34:909-17.
Husain S, Alexander BD, Munoz P, Avery RK, Houston S, Pruett T. Opportunistic mycelial fungal infections in organ transplant recipients: emerging importance of non-Aspergillus mycelial fungi. Clin Infect Dis. 2003; 37:221-9.
Friedman DZP, Schwartz IS. Emerging Fungal Infections: New Patients, New Patterns, and New Pathogens. J Fungi. 2019; 5: 67-85.
Jayaprada R, Nagaraja M, Kumar GLS, Venkataramana B, Kalawat U. Common fungal isolates from routine clinical specimens - A two-year study from a tertiary care hospital in South India. J Clin Sci Res. 2017; 6:2-9.
Swamy MK, Akhar MS, Sinniah UR. Antimicrobial Properties of Plant Essential Oils against Human Pathogens and Their Mode of Action: An Updated Review. Evid Based Complement Alternat Med. 2016; 3012462.
Leaw SN, Chang HC, Sun HF. Identification of medically important yeast species by sequence analysis of the internal transcribed spacer regions. J Clin Microbiol. 2006; 44: 693-9.
Inouye S. Laboratory evaluation of gaseous essential oils (part 1). Int J Aromather. 2003; 13:95-107
Sukatta U, Haruthaithanasan V, Chantarapanony W, Suppakul P. Antifungal activity of cinnamon oil and their synergistic against postharvest decay fungi of grape in vitro. Kasetsart J Nat Sci. 2008; 42(5):169-74.
Eman-abdeen E, El-Diasty EM. Antifungal activity of clove oil on dermatophytes and other Fungi. Int J Adv Res. 2015; 3(12):1299 - 305.
Pina-Vaz C, Gonçalves Rodrigues A, Pinto E, Costa-de-Oliveira S, Tavares C, Salgueiro L. Antifungal activity of Thymus oils and their major compounds. J Eur Acad Dermatol Venereol. 2004; 18(1):73-8.
Kumar A, Thakur S, Thakur VC, Kumar A, Patil S, Vohra MP. Antifungal activity of some natural essential oils against Candida species isolated from blood stream infection. JKIMSU. 2012; 1(1):61-6.