# Antimicrobial Efficacy of Edible Mushroom Extracts: Assessment of Fungal Resistance
## 3 Results and Discussion
### Antifungal Activity of T. camphoratus Methanol Extract (TcM)
#### 3.2.2 Antifungal Activity Test of TcM on Apple or Grape JA (Complex Medium)
This MAPK pathway is responsive to oxidative (osmotic) cues, and hence protects the wild type fungal cells from environmental oxidative (osmotic) stressors. \begin{table}
\begin{tabular}{c c c c} \hline \hline
**Extract (\%; _wt/v_)** & **0.0** & **0.8 1** & **1.6 1** \\
**Fungi** & & & \\ \hline
**Apple JA:** & & & \\ _N. fischer_ 96468 & 100 \(\pm\) 0\% & 32 \(\pm\) 0\% & 15 \(\pm\) 2\% \\ _A. flavus_ 3357 & 100 \(\pm\) 3\% & 66 \(\pm\) 0\% & 41 \(\pm\) 6\% \\ _A. parasitic_ 5862 & 100 \(\pm\) 9\% & 57 \(\pm\) 3\% & 29 \(\pm\) 12\% \\ _P. expansum_ W1 & 100 \(\pm\) 0\% & 45 \(\pm\) 12\% & 32 \(\pm\) 19\% \\ _P. expansum_ FR2 & 100 \(\pm\) 0\% & 47 \(\pm\) 30\% (\(p\) \(<\) 0.5) 2 & 40 \(\pm\) 0\% 2 \\ _P. expansum_ W2 & 100 \(\pm\) 7\% & 56 \(\pm\) 5\% & 31 \(\pm\) 37\% \\ _P. expansum_ FR3 & 100 \(\pm\) 0\% & 41 \(\pm\) 40\% (\(p\) \(<\) 1) 2 & 4 \(\pm\) 100 \(\pm\) 34\% (\(p\) \(=\) 1) 2 \\ \hline Average & 100 \(\pm\) 3\% & 65 \(\pm\) 13\% & 41 \(\pm\) 16\% \\ \hline
**Grap JA:** & & & \\ _N. fischer_ 96468 & 100 \(\pm\) 6\% & 45 \(\pm\) 9\% & 41 \(\pm\) 3\% \\ _A. flavus_ 3357 & 100 \(\pm\) 6\% & 66 \(\pm\) 11\% (\(p\) \(<\) 0.01) & 55 \(\pm\) 9\% \\ _A. parasticus_ 5862 & 100 \(\pm\) 5\% & 58 \(\pm\) 10\% & 58 \(\pm\) 3\% \\ _P. expansum_ W1 & 100 \(\pm\) 10\% & 56 \(\pm\) 10\% & 43 \(\pm\) 8\% \\ _P. expansum_ FR2 & 100 \(\pm\) 17\% & 49 \(\pm\) 40\% (\(p\) \(=\) 0.8) 2 & 4 \(\pm\) 100 \(\pm\) 28\% (\(p\) \(=\) 1) 2 \\ _P. expansum_ W2 & 100 \(\pm\) 11\% & 53 \(\pm\) 6\% & 40 \(\pm\) 8\% \\ _P. expansum_ FR3 & 100 \(\pm\) 16\% & 4 \(\pm\) 40\% (\(p\) \(<\) 0.5) 2 & 4 \(\pm\) 108 \(\pm\) 43\% (\(p\) \(<\) 1) 2 \\ \hline Average & 100 \(\pm\) 10\% & 75 \(\pm\) 18\% & 64 \(\pm\) 15\% \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Antifungal activity of TcM on apple or grape JA (% radial growth compared to no treatment control). However, fungi with mutations in this MAPK system are not affected by the toxicity of FLU, which results in the development of FLU tolerance in the environment [49]. As a proof of concept, we examined the susceptibility of the wild type and MAPK mutants of _A. fumigatus_ (_sakA\(\Delta\)_ and _mpkC\(\Delta\)_) and of _P. expansum_ (FR2) (Table 1) [36; 37; 38] to a benzoic compound SA, a natural aromatic constituent found in many plant species [56]. In a prior study, aromatic hydrocarbons such as pentachloronitro-benzene also exerted its antifungal activity via an action similar to FLU [50]. SA is a redox-active benzoic derivative, possessing both antioxidant and prooxidant potential [57; 58; 59]. Therefore, we reasoned that SA exerts its antifungal activity by interfering with fungal antioxidant systems such as the MAPK pathway, whereby the wild type fungus will be sensitive to the treatment while the antioxidant MAPK mutants, as shown in this study, will be tolerant or less susceptible. According to its vapor pressure value (5.93 \(\times\) 10\({}^{-1}\) mm Hg, 25 \({}^{\circ}\)C), SA exists mainly as a vapor in the environment [60]. To target fungi, SA was remotely applied from the SA-saturated membrane filter placed on each plate. As shown in Figure 2, the growth of the wild type of _A. fumigatus_ (AF293) or _P. expansum_ (W1) was completely inhibited by FLU (50 \(\upmu\)M), while MAPK mutants (_sakA\(\Delta\)_, _mpkC\(\Delta\)_, FR2) of both species exhibited tolerance to FLU. Fungal MAPK mutants were also discovered to be tolerant to the natural benzoic SA (Figure 2). For example, _A. fumigatus_sakA\(\Delta\)_ and _mpkC\(\Delta\)_ mutants showed higher tolerance (higher radial growth) to FLU or SA compared to the wild type AF293, which showed almost no growth under the same treatment. Similar trends were also observed in the _P. expansum_ FR2 mutant when compared to the wild type W1. Results indicate that, similar to FLU, SA also acts as an inducer of \"oxidative (osmotic) imbalance\" in fungal cells, thus indicating that both SA and FLU might share a common mechanism of antifungal action.