## 4 Discussion
Therefore, FAPROTAX database still needs to be extended to cover other relevant functions in soils. This study provided soil bacterial taxa capable of nitrogen fixation, phosphate and potassium solubilization, so further study on soil sample could make the most of our work. Although this study has demonstrated that FAPROTAX was compatible with soil samples, the bias of this tool should be kept in mind. The FAPROTAX has an assumption that if all cultured members of a taxon can perform a function, all members of that taxon (cultured and uncultured) can perform that function. Moreover, since soil ecosystem contains several biogeochemical cycles as well as diverse prokaryotic taxa, it is concerning that FAPROTAX may have low performance (low functional assignment percentage) for certain soil samples. We recommend that further work should add more soil-specific functions and prokaryotic taxa to optimize the performance of this tool. On the other hand, other alternative tools can be applied for bacterial functional annotation in soil, such as PICRUST([PERSON] and others, 2018; [PERSON] and others, 2018) or Tax4 Fun ([PERSON] and others, 2018). Notice should be taken that each tool provides different aspects of bacterial functional phenotypes. FAPROTAX presents functional phenotypes as metabolic and ecologically relevant functions which are predicted based on the literature of cultured taxa (i.e., nitrogen fixation, nitrification, hydrocarbon degradation, chitinolysis, cellulolysis, etc. ), whereas PICRUST or Tax4 Fun present functions as phenotypes of gene families or enzyme activities which are predicted based on gene content. We simulate results from Tax4 Fun to show the functional phenotype of bacteria associated with the rhizosphere soil samples of _Trifolium pratense_ used in this study (Appendix A). The simulated results show various potential enzymes that may be relevant to many soil functions (Figure A2). In our opinion, each functional annotation tool provides information on different aspects of bacterial functions. Selection of such tools for a particular study should depend on its purposes. If a study focuses on key bacterial functions important for biogeochemical cycling as well as microbec microbe, plant-microbe and animal-microbe interactions, FAPROTAX can be a suitable choice. On the other hand, if a study targets at changes of gene expression or potential enzyme activity, other tools should be considered, such as PICRUST or Tax4 Fun. In conclusion, this study presented that FAPROTAX database can be effectively used to predict function of bacteria in soil samples. Even though the database cannot predict function of all detected taxa, it can be beneficial for fast-functional screening or grouping of 16S derived bacterial data from any ecosystem. We further suggested that additional datasets of both the taxonomy and functional references could improve the FAPROTAX database and thereby increase the number of functionally assigned OTUs derived from 16S rRNA. The following are available online at [[https://www.mdpi.com/2076-3417/11/2/688/s1](https://www.mdpi.com/2076-3417/11/2/688/s1)]([https://www.mdpi.com/2076-3417/11/2/688/s1](https://www.mdpi.com/2076-3417/11/2/688/s1)); Figure S1: Heatmap of all detected functions of bacteria across all soil samples. The data based on OTUs occurrence (number of OTUs capable for each function) derived from agricultural bulk soil (TB), agricultural rhizosphere soil (TRh), forest soil (SF) and mangrove-rhizosphere soil (Rs) samples. Table S1.1: OTU table of prokaryotic taxa detected in agricultural soil. Table S1.2: OTU table of prokaryotic taxa detected in forest soil. Table S1.3: OTU table of prokaryotic taxa detected in mangrove-rhizosphere soil. Table S2.1: FAPROTAX validation of prokaryotic taxa prokaryotic taxa detected in agricultural soils showing evident based on peer-reviewed literature of soil ecosystems on the appearance and the functional performance of prokaryotic taxa detected in agricultural soils.