## 2 Materials and Methods
### 165 rRNA Gene Analysis
#### 2.3.1 DNA Extraction, PCR Amplification, and 165 rRNA Gene Sequencing Analysis
\begin{table}
\begin{tabular}{l l c c c c c c c c c c c c} \hline \hline
**Field** & **Field Size** & **Lotus** & **Damage Index** & **pH** & **EC** & **NH\({}_{4}\)**-N** & **P\({}_{2}\)O\({}_{5}\)** & **K\({}_{4}\)O** & **CaO** & **MgO** & **SG\({}^{*}\) & **TN** & **TC** & **C/N** & **CEC** \\ \hline
**Name** & **(m\({}^{3}\))** & **Cultivar** & **(0,1,2,3,4)** & **(-)** & **(dS/m)** & & **(mg/100 g-Dry Soil)** & & **(-)** & **(+)** & **(+)** & **(+)** & **(+)** & **(+)** & **(+)** & **(+)** & **(+)** & **(**-)** \\ \hline
1 A\({}^{1}\)-A & 1e. & 0000 & 2.42 \(\pm\) 0.99 (14) & 5.8 & 0.44 & 5.3 & 34.8 & 28.8 & 383.6 & 110.2 & 0.39 & 4.88 & 41.0 & 10.1 & 27.5 \\ LA\({}^{2}\) & ca. & 1000 & \"Koshuku\" & 2.60 \(\pm\) 0.82 (25) & 6.0 & 0.55 & 39.3 & 38.1 & 66.2 & 513.0 & 146.5 & 0.30 & 0.62 & 61.7 & 9.9 & 39.3 \\ IB-1 & ca. & 1000 & \"Koshuku\" & 2.34 \(\pm\) 1.29 (25) & 6.0 & 0.56 & 13.7 & 58.9 & 54.6 & 418.8 & 133.1 & 0.43 & 0.49 & 5.03 & 10.3 & 26.1 \\ IB-2 & ca. & 1000 & \"2.07 \(\pm\) 0.83 (27) & 6.1 & 0.70 & 22.2 & 52.8 & 65.7 & 532.8 & 187.6 & 0.31 & 0.67 & 6.80 & 10.1 & 36.9 \\ IC-1 & ca. & 1100 & \"Kanasumi\" & (020) & 6.3 & 0.18 & 20.7 & 10.2 & 50.1 & 34.90 & 73.4 & 0.68 & 3.89 & 3.94 & 10.4 & 24.0 \\ IC-2 & ca. & 1100 & No. 24\" & 0 (19) & 6.8 & 0.14 & 23.4 & 23.9 & 65.4 & 278.0 & 71.2 & 1.04 & 0.35 & 2.88 & 8.3 & 18.6 \\ ID-1 & ca. & 1000 & \"Power\" & 2.41 \(\pm\) 0.94 (17) & 6.7 & 0.59 & 68.1 & 71.4 & 66.5 & 353.1 & 10.3 & 0.32 & 0.66 & 6.53 & 10.0 & 36.8 \\ ID-2 & ca. & 1000 & \"Power\" & 2.42 \(\pm\) 0.90 (12) & 6.5 & 0.42 & 21.1 & 59.0 & 47.8 & 533.1 & 99.5 & 0.48 & 0.56 & 5.71 & 10.2 & 34.3 \\ IE-1 & ca. & 1000 & \"Koshuku\" & 1.75 \(\pm\) 0.79 (20) & 5.8 & 0.47 & 13.7 & 29.4 & 42.1 & 394.0 & 69.4 & 0.48 & 0.48 & 4.96 & 10.3 & 27.6 \\ IE-2 & ca. & 1000 & \"Koshuku\" & 1.52 \(\pm\) 0.64 (27) & 5.6 & 0.51 & 12.0 & 33.0 & 33.5 & 35.83 & 61.6 & 0.57 & 0.47 & 4.9 & 24.9 & 24.9 \\ IF-1 & ca. & 1000 & \"Koshuku\" & 1.18 \(\pm\) 0.90 (28) & 6.0 & 0.28 & 9.6 & 33.3 & 46.0 & 302.4 & 53.4 & 0.67 & 0.39 & 4.01 & 10.3 & 21.4 \\ IE-2 & ca. & 1000 & \"Koshuku\" & 1.90 \(\pm\) 0.91 (20) & 5.9 & 0.53 & 17.8 & 33.4 & 47.7 & 395.9 & 71.9 & 0.47 & 0.48 & 5.00 & 10.3 & 28.9 \\ IG & ca. & 2000 & \"Power\" & 2.67 \(\pm\) 0.58 (3) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) \\ HI & ca. & 1000 & \"Matsukaze\" & 3.25 \(\pm\) 0.50 (4) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) \\ NI & ca. & 4000 & \"Damu\" & 3.25 \(\pm\) 0.47 (20) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) & \(-\) \\ \hline \hline \end{tabular}  
* The first letter indicates the location of the fields. I: Ibaraki and N: Niigata. \({}^{\rm b}\) The number indicates the duplicate samples from the different locations in the field. \({}^{\rm c}\) Specific gravity. \({}^{\rm d}\) Number of lotus tubers used for damage index evaluation. \({}^{\rm e}\) Not determined.  
\end{table} 
Table 1: Summary of lots cultivation fields, damage index values of lotus tubers, and soil chemical analysis data. the DADA2 treatment, the sequences were clustered at \(\geq\)97% similarity into operational taxonomic units (OTUs) using VSEARCH [18]. Taxonomic assignment was performed using classify-sklearn retained on the SILVA database version 138 [19]. Representative OTUs were chosen on the basis of the \(>\)1% average abundance rate.