## 3 Results
### The Effect of Plasma on the Genomic DNA of _A. oryzae_  
To determine the plasma-induced changes in the genomic DNA of _A. oryzae_, whole genome sequencing was performed on fungal samples cultured for 48 h after the spores in PDB were treated with an NTAPP jet for 5 min (short time) and 20 min (long time) or left untreated (control). The genome size of _A. oryzae_ is \(\sim\)37 Mb and contains 12,074 genes on 8 chromosomes [27; 28]. In our analysis, only variations observed in most reads were considered mutations.  
Figure 5: Oxidation reduction potential (ORP) measurements in the PDB media post plasma treatment for 5, 10, 15, and 20 min, or without treatment (0 min). Post plasma treatment, the fungal spores in the PDB solution were cultured for 168 h, and the ORP was measured every 24 h. Each value is indicated as the mean and standard deviation of 9 replicate measurements.  
Figure 6: The pH measurement in the PDB media post plasma treatment of the fungal spores for 5, 10, 15, and 20 min. Untreated (0 min) fungal spores were used as a control. The pH was measured immediately after plasma treatment. Each value is indicated as the mean and standard deviation of 9 replicate measurements.  
Compared to the control DNA (no plasma treatment), the genomic DNA from the plasma-treated samples (5 min and 20 min treatments) displayed approximately 0.0016-1.0017% variations in sequence (single base pair changes, deletions, and insertions) (Table 2 and Supplementary Tables S1 and S2). No dramatic differences in sequence were observed between the samples treated with plasma for 5 min and 20 min, although a slightly higher mutation rate was exhibited in the samples treated for 5 min (0.0017%) than in those treated for 20 min (0.0016%) (Table 2). The total mutation percentage in the exon area was about 0.0001-0.0002% in the samples treated with plasma for 5 min and 20 min, which was lower than that in the intron area (Table 2). Missense mutations (changes in the amino acid sequence of a gene) occurred at approximately 0.0004-0.0001% positions in 17 and 10 genes upon plasma treatment for 5 min and 20 min, respectively (Table 2 and Supplementary Table S3).  
Variations in the genomic DNA appeared on each chromosomal DNA with a similar proportion (Table 3 and Supplementary Tables S1 and S2). The percentage of mutations was slightly higher in chromosome 4 than in the other chromosomes. No change was observed in the mitochondrial DNA (Table 3 and Supplementary Tables S1 and S2).  
\begin{table}
\begin{tabular}{c c c c} \hline \hline  & **Treatment Time (min)** & **Mutation Position Number** & **Mutation (\%)** \\ \hline \multirow{2}{*}{**Total mutations**} & 5 & 655 & 0.0017 \\  & 20 & 596 & 0.0016 \\ \hline \multirow{2}{*}{**Total mutations in exons**} & 5 & 62 & 0.0002 \\  & 20 & 25 & 0.0001 \\ \hline \multirow{2}{*}{**Total mutations in introns**} & 5 & 593 & 0.0016 \\  & 20 & 571 & 0.0015 \\ \hline \multirow{2}{*}{**Total missense mutations**} & 5 & 35 (17 1)  & 0.0001 (0.1377 2  ) \\  & 20 & 17 (10 1)  & 0.0004 (0.0810 2  \\ \hline \hline \end{tabular}
\end{table}
Table 2: Total mutations in the genomic DNA of _A. oryzae_ after treatment with plasma for 5 min and 20 min.  
Figure 7: The mRNA levels of two putative genes involved in vesicle trafficking and transport in fungal hyphae cultured for 48 h post plasma treatment of the spores. Each value is indicated as the mean and standard deviation of 6 replicate measurements. **_p_ < 0.01 and **_p_ < 0.05.