Comparison of Broth Dilution and Disc Diffusion Method for the Antifungal Susceptibility Testing of Aspergillus flavus
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With the discovery of newer antifungal agents, emergence of resistance among Aspergillus, is a major problem.In the present study, 25 different clinical isolates of Aspergillus flavus were screened for their susceptibility against five common antifungal agents by broth dilution and disc diffusion method with the aim to evaluate both the methods for their suitability to adapt in clinical laboratory.All the isolates showed susceptibility to five antifungals by broth dilution after 24 hrs however all isolates showed resistance for 5FC, fluconazole and ketoconazole after 48 hrs.By disc diffusion method, all isolates were sensitive for all antifungals except fluconazole which showed resistance even after 24 hrs.Amphotericin B and itraconazole were in good agreement between both the methods where 100% isolates were sensitive at both time period by broth dilution and 84% to 100% by disc diffusion method.Both the methods are in good agreement and since disc diffusion method is easy to perform, cheap, rapid and same can be adapted easily for the routine susceptibility testing of clinical fungal isolates in medical microbiology laboratory.Keywords:
Dilution
The essential oil of Ammodaucus leucotrichus Coss. & Dur. seeds growing wild in SouthWest of Algeria were obtained by hydrodistillation and analysed by GC and GC-MS. Other parameters such as density, refractive index, optical rotation, freezing point, solubility in ethanol, acid value and ester index are also measured. The antifungal properties of the essential oil were conducted using biomass technique on liquid medium, mycotoxigenic test, mycelial growth, determination of minimum inhibitory concentration (MIC), sporulation and germination spores’. The essential oil was found to be active against Aspergillus flavus MTTC 2799 (Microbial Type Culture Collection and Gene Bank) and Aspergillus ochraceus CECT 2092 (Spanish Type Culture Collection). The GC and GC-MS data showed a total of 19 compounds were identified in the hydrodistilled oil. The oil was dominated by oxygenated monoteprenes represented by perilla aldehyde (81.62%).
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Aspergillus terreus
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Summary The red algae A sparagopsis taxiformis collected from the S traits of M essina ( I taly) were screened for antifungal activity against A spergillus species. EUCAST methodology was applied and extracts showed antifungal activity against A . fumigatus , A . terreus and A . flavus . The lowest minimum inhibitory concentrations observed were <0.15 mg ml −1 and the highest were >5 mg ml −1 for A spergillus spp. tested. Agar diffusion assays confirmed antifungal activity of A . taxiformis extracts in A spergillus species.
Aspergillus terreus
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Different bacteria and fungi live as commensal organisms as part of the human microbiota, but shifts to a pathogenic state potentially leading to septic infections commonly occur in immunocompromised individuals. Several studies have reported synergistic or antagonistic interactions between individual bacteria and fungi which might be of clinical relevance. Here, we present first evidence for the interaction between Klebsiella pneumoniae and several Aspergillus species including A. fumigatus, A. terreus, A. niger and A. flavus which cohabit in the lungs and the intestines. Microbiological and molecular methods were employed to investigate the interaction in vitro, and the results indicate that Klebsiella pneumoniae is able to prevent Aspergillus spp. spore germination and hyphal development. The inhibitory effect is reversible, as demonstrated by growth recovery of Aspergillus spp. upon inhibition or elimination of the bacteria, and is apparently dependent on the physical interaction with metabolically active bacteria. Molecular analysis of Klebsiella-Aspergillus interaction has shown upregulation of Aspergillus cell wall-related genes and downregulation of hyphae-related genes, suggesting that Klebsiella induces cell wall stress response mechanisms and suppresses filamentous growth. Characterization of polymicrobial interactions may provide the basis for improved clinical management of mixed infections by setting the stage for appropriate diagnostics and ultimately for optimized treatment strategies.
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Dereplication of the antifungal extracts of Aspergillus flavus indicated that the primary antifungal compound present was the known aspirochlorine (1). Preparative isolation work resulted in the identification of the new compounds tetrathioaspirochlorine (2) and cyclo(d-N-methyl-Leu-l-Trp) (3).
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The fumigation treatment with oregano essential oil (OEO) was applied to inactivate Aspergillus flavus (A. flavus) and its mechanism was studied. Results indicated that the minimal fungicidal concentration (MFC) of OEO vapor was lower than OEO dissolved in liquid medium. Indeed, OEO vapor at MFC resulted in the abnormal branching of A. flavus mycelium. Further, the fumigation time at MFC required for fungicidal effect was reduced from 7 days to 8 h by enhancing the fumigation temperature from 28 to 45°C. Meanwhile, the mechanism of action might be the oxidative stress in response to OEO vapor within the cells of A. flavus mycelium. Results suggested increased lipid peroxidation and O2− production as well as SOD/CAT activity, eventually leading to decreased mitochondrial activity and ATP production. Additionally, OEO vapor inhibited metabolic activities related to the synthesis of protein and methylglyoxal as well as aflatoxin production. Novelty impact statement OEO exhibited a higher fungicidal effect on A. flavus mycelium compared to its spores. OEO fumigation exhibited an inhibitory effect on the metabolic synthesis of A. flavus. OEO fumigation induced oxidative stress and decreased mitochondrial activity.
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Evaluation of antifungal activity of some oils from Algerian medicinal plants against Aspergillus flavus strain produced aflatoxinsA. Amrouche, H. Benmehdi, A. Moussaoui, K. Mebarki, A Chaoufi, A. Saneba, A. Lazouni .H2, D. Chabane Sari
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