Aflatoxin production and in vitro toxicity of Aspergilli section Flavi isolated from air samples collected from different environments

Abstract

Aspergilli section Flavi, originally isolated from air samples collected from inhabited apartments (AP), unoccupied basements (BS), and processing facilities of a grain mill (GM), were analyzed for their potential to produce aflatoxin B 1 (AFB 1) on solid media. The isolates were further characterized with regard to their cytotoxic, genotoxic, and pro-inflammatory properties in vitro. Aspergilli were identified based on partial calmodulin (CaM) gene sequencing; the producing capacities of isolates were analyzed by HPLC/FLD and confirmed by genes in biosynthesis (aflR, norA, omtA). In the grain mill, the Aspergilli section Flavi (up to 1.3 × 10 6 cfu/m 3) dominated by AFB 1-producing Aspergillus flavus (71%, 4.5-5254 ng/ml) which showed a serious health risk for workers. Living environments were not relevant sources of exposure. After 24 h, AFB 1 (1-100 μmol/l) reduced cell viability (MTT test) in both A549 cells and THP-1 macrophage-like cells without reaching IC 50. In A549 cells, the extract of the AFB 1producing A. flavus significantly decreased cell viability but not below 50%. THP-1 macrophage-like cells were more sensitive to both extracts, but IC 50 was obtained only for the AFB 1-producing strain (0.37 mg/ml; AFB 1 2.78 μmol/l). AFB 1 (1 and 10 μmol/ l) induced significant DNA damage (tail intensity, alkaline comet assay) in A549 cells in contrast to Aspergilli extracts. AFB 1 elevated IL-6 and IL-8, while Aspergilli extracts increased IL-1β, TNF-α, and IL-17 release in THP-1 macrophages (ELISA). Chronic exposure to AFB 1 and/or other metabolites in airborne A. flavus from occupational environments may stimulate epithelial damage of airways accompanied by lowered macrophage viability.

Environment Changes, Aflatoxins, and Health Issues, a Review

“…Production of these compounds is highly susceptible to environmental factors, pre-and/or post-harvest, thus, when changes in the weather occur, mycotoxins will be affected [4]. The situation is even more complicated by the fact that contamination also occurs during storage and transportation because different indoor environments are recognised sources of both fungal producers and their metabolites [6][7][8][9][10].…”

“…Although these fungal species are distributed worldwide, AFs producers were always more prevalent in tropical and subtropical regions. Based on recent studies, AFs produce fungi and consequently AFs are becoming more prevalent in countries with temperate climate [3,[10][11][12][13][14].…”

Combined Toxicity of the Most Common Indoor Aspergilli

“…Human lung adenocarcinoma cells A549 and human monocytic leukemia cells THP-1 (European Collection of Cell Cultures, Salisbury, UK) were grown at 37 °C in 5% CO 2 in 75 cm 2 flasks in RPMI supplemented with glutamine (2 mmol/L), heat-inactivated FBS 10%, penicillin (100 IU/mL; 1 IU 67.7 μg/mL) and streptomycin (100 μg/mL). Extracted Aspergilli were prepared as published previously [ 12 , 17 , 37 ] following the microextraction procedure according to Smedsgaard et al [ 38 ]. Weighed dried fungal extracts were dissolved in 100% DMSO.…”

“…The first criterion was that the concentrations of choice had non-significant effect on either A549 or THP-1 macrophage-like cell viability. Each extract was previously tested on most significant mycotoxins, i.e., AFB 1 for the section Flavi [ 12 ], FB 2 for the series Nigri [ 37 ] and STC for the series Versicolores [ 17 ]. Thus, the second criterion was to combine the mycotoxin producers from the section Flavi or the series Versicolores with the mycotoxin-producing isolates from the series Nigri and mycotoxin non-producing isolates from the sections Flavi or series Versicolores with the series Nigri .…”

“…Because of their biomedical and economical significance, special importance among indoor Aspergilli is given to those from the section Flavi and series Nigri (ex-section Nigri ) and Versicolores (ex-section Versicolores ), among which A. flavus , A. tubingensis , A. welwitschiae , A. jensenii and A. creber are the most commonly identified species [ 11 , 12 , 13 , 14 , 15 , 16 , 17 ]. The ability to tolerate a range of the water activities (a w ) 0.75–0.90, a wide range of pH and ionic strength make Aspergilli particularly common indoor fungi [ 18 ].…”