Obtaining monokaryotic and dikaryotic mycelial cultures of two Amazonian strains of Geastrum (Geastraceae, Basidiomycota)

Abstract

The high diversity of the genus Geastrum and the difficulty of obtaining mycelial cultures impairs the study of the ecophysiology and the exploration of the biotechnological potential of the taxon. In this study, different culture media were tested to obtain mycelial cultures for G. lloydianum and G. subiculosum collected in the Brazilian Amazon. Data on spore germination, and isolation of monokaryotic cultures and in vitro sexual reproduction are presented, as well as a brief morphological description of the cultures obtained. For both species, Potato Dextrose Agar (PDA) was the most promising of the tested culture media. The highest growth in agar culture ever recorded for this genus is reported (4.9 mm per week for G. lloydianum and 7.5 mm for G. subiculosum). In the PDA culture medium, spores germinated after 35-40 days of incubation and the isolation of monokaryotic cultures of the two species, as well as in vitro sexual crosses, were successfully performed.

Biodegradation and reduction of toxicity of Azo Trypan Blue dye by Amazonian strains of gasteroid fungi (Basidiomycota)

“…After growth, fragments of 3 × 3 mm from the edge of the cultures were inoculated in the center of new Petri dishes with equal volume and content. Then they were incubated under the conditions suggested by Santana et al (2020b) to obtain the pure culture. Thus, each species was considered a treatment, totaling six treatments.…”

“…For the discoloration test of the blue trypan dye in solid medium, a 3 × 3 mm block was used, containing mycelium removed from the edge of the pure strain of each species and transferred to the centre of a Petri dish (90 mm in diameter) containing 15 mL of the homogeneous mixture of PDA plus trypan blue (0.05%). Petri dishes were incubated according to the indications of Santana et al (2020b), and each plate was considered a replicate. Every three days, from the beginning of the experiment, for a period of 21 d of incubation, the dye discoloration halo was evaluated by measuring the diameter with a ruler (orthogonal directions).…”

“…The mixture was homogenized and sterilized in autoclave at 121 °C for 15 min. The vials were incubated at 25 ± 2 °C according to the indications for cultivation of Santana et al (2020b), and each bottle was considered a replicate. Dye discoloration was analyzed every 20 days from the beginning of the experiment, for a period of 60 days.…”

“…Culture medium supplementation (DP) was used because it made discoloration more efficient (Chang et al, 2000;Bardi and Marzona, 2010), since azo dyes, in general, have carbon and nitrogen deficiency (Stolz, 2001), elements of great relevance in the development of fungal cultures. Furthermore, supplementation has positive results in the development of Geastrum strains (Santana et al, 2020b).…”

“…Although the lacase enzyme is likely responsible for degrading trypan blue dye, as it was for other synthetic dyes, studies should consider that fungi secrete several other enzymes that together can be even more efficient in the degradation of this dye than purified enzymes (Camarero et al, 2005). The combined effect of the enzymatic system of gasteroid fungi, combined with the cultivation conditions to which the strains were submitted (Santana et al, 2020b), may have occurred in this study, resulting in the different outcomes of the degradation and toxicity of the trypan blue dye. It is possible that other variables, such as supplementation of culture medium, pH, temperature and agitation, may increase or accelerate the enzymatic activity of these species, as reported for other fungi (Asgher et al, 2008;Anastasi et al, 2010;Bibi and Bhatti, 2012).…”