Selective Flamingo Medium for the Isolation of Aspergillus fumigatus

Abstract

For various studies in the clinic as well as the environment, it is essential to be able to selectively isolate Aspergillus fumigatus from samples containing bacteria as well as various other fungi (mainly Mucorales). Six agar media were compared for effectiveness in selectively isolating Aspergillus fumigatus from agricultural plant waste, woodchip waste, green waste, soil, grass and air samples collected in the Netherlands at a 48 °C incubation. The Flamingo Medium incubated at 48 °C, provided the most effective condition for the isolation of A. fumigatus from environmental samples, since it effectively inhibited the growth of competing fungi (mainly Mucorales) present in the environmental samples. Flamingo Medium reduced the number of colonies of Mucorales species by 95% and recovered an average of 20−30% more A. fumigatus colonies compared to the other media. We further confirmed that Flamingo Medium can inhibit the growth of clinical Mucorales, which occasionally present in patient’s tissue and can also be used for clinical applications. We suggest the use of Flamingo Medium as an efficient method for the study of A. fumigatus from important environmental niches for which there is increasing interest. Additionally, it can also be used in the clinic to isolate A. fumigatus especially from tissue contaminated with Mucorales.

Catching more air: A method to spatially quantify aerial triazole resistance inAspergillus fumigatus

“…These colonies could however still be readily identified as A. fumigatus by their compact and circular morphology. This was because few fungi other than A. fumigatus and morphologically distinct Mucorales species grew on the medium, as detailed in Zhang et al 2021. Figure 3: Visual examples of A. fumigatus growth on the sticky seals covered with Flamingo medium. Plates of most outdoor samples showed some Mucorales growth but not to a degree that hindered CFU counts.…”

“…Under sterile conditions, the covers were removed from the seals and then individually placed in square Petri dishes with vents (120mmX120mmX17mm), sticky-side-up, again only touching the non-sticky strip with clean hands. Once all seals were placed in this manner, flamingo medium (Zhang et al 2021) was poured into the plate such that the entire seal and all of the plate was covered with a thin layer of medium. Crucially, the medium was prepared as described in (Zhang et al 2021) except that the medium, after the autoclaving step, was allowed to cool overnight in an incubator set at 60°C before adding the antimicrobial supplements and pouring the medium.…”

“…The isolation of A. fumigatus resistance genotypes from plant waste material and soils that could be linked to those found in patients has been instrumental in establishing the environment as a source for triazole resistance. Crucially the development of selective media such as the flamingo type (Zhang et al 2021) has not just allowed for the isolation, but also accurate counts of A. fumigatus colony forming units (CFU). Plating aliquots of a sample on a selective medium with and without different types of triazoles has enabled researchers to not just qualitatively but also quantitively assess resistance by determining the resistance fractions.…”

“…However, prolonged exposure also means potential increased exposure to other microbes. Here we tested a culturing method where we used the highly selective Flamingo medium (Zhang et al 2021) by pouring it directly on top of exposed seals. Besides exposing sticky seals indoors, we also tested whether the use of foldable delta traps could be combined with the use of the sticky seals in a flexible, effective protocol for cheap and large-scale outdoor spore capture, including citizen-science initiatives.…”

Aspergillus and Health

“…Culture remains the gold standard for the laboratory diagnosis of invasive fungal infections, including aspergillosis. Aiming to attain a higher sensitivity of cultures (especially in samples contaminated with different fungal genera), Zhang et al [ 1 ] suggested the use of Flamingo Medium as an efficient method for the isolating A. fumigatus , especially from tissue contaminated with fungi belonging to Mucorales order. The authors reported that, using Flamingo Medium, they were able to recover an average of 20−30% more A. fumigatus colonies compared to other media, and at the same time, a 95% reduction in the number of Mucorales colonies was also observed.…”