## 3 Results and Discussion
### Removal Efficiency  
The potential of the 2 De strain to reduce the Mn concentration was investigated. During this process, bacteria can remove metals from soil by both bioaccumulation and biosorption [39]. It was reported that living cells have a greater capacity for metal removal than non-living cells [40]; therefore, live bacterial cells were used in the experiment. The pH value and surface tension were carefully controlled over the entire period of bacteria cell and soil contact. When soil concentration increased from 1% to 6%, Mn removal efficiency increased from 46% to 81% after three days of incubation (Figure 3). After six days, the Mn removal efficiencies in all research samples were comparable and were within the range of 85-89%. At the end of the experiment, the highest Mn removal (88%) was recorded in the set containing soil at a concentration (_w/v_) of 4%, but it was not significantly higher compared to the sets enriched in 2% and 6% (_w/v_) soil. The high percentage of Mn accumulation can be attributed to the fact that the 2 De strain has been isolated from the soil from metallurgical waste heap. In such an environment, autochthonous strains demonstrate high tolerance to excess heavy metal concentration due to the stress coping mechanisms. Some of these mechanisms, especially utilizing proteins, are certainly present in 2 De cells, as shown in the SDS/PAGE analysis (Figure 2). The results obtained in this study showed that throughout the duration of the experiment the pH increased (Figure 4a), and this certainly affects the increase in Mn removal efficiency (Figure 3). Figure 3: Manganese removal efficiencies of the 2 De strain after incubation with different concentrations of soil at 28 \({}^{\circ}\)C for 18 days. The same letter indicates no significant differences among groups (LSD, _p_\(<\) 0.05). Error bars are the standard deviation (_n_\(=\) 3). Figure 4: Variation in pH (**a**) and in surface tension (**b**) leachate observed during the accumulation study of Mn by the 2 De strain using different soil concentration from the metallurgical waste. The same letter indicates no significant differences among groups (LSD, \(p<0.05\)). Error bars are standard deviation (\(n=3\)). These results suggested that strain 2 De can self-mediate pH changes of medium K in whole cultivation period. This self-mediated pH of 2 De strain is an interesting phenomenon, which is worthy further investigation to learn how this phenomenon affects biosorption and bioaccumulation of Mn by the growing 2 De cells. Our results were well agreed with the findings of [PERSON] et al. [41] who revealed that _Bacillus cereus_ strain Cd01-mediated pH decrease of the cultivation medium resulting in inhibition of Cd\({}^{2+}\) biosorption of on Cd01 cells at the initial cultivation period. Along with the duration of the cultivation period an increased pH was observed, which was associated with facilitated biosorption of Cd\({}^{2+}\) on Cd01 cells. [PERSON] et al. [42] noted that high pH (<7.5) promoted biosorption of metal on cell surface and its bioaccumulation inside cell due to the deprotonation of metal binding sites on cell surface. Other researches observed the low Mn\({}^{2+}\) adsorption rate in an acidic condition (pH < 5), caused by the large amounts of protons like H\({}_{3}\)O\({}^{+}\) and H\({}^{+}\), which increased the difficulty in binding sites of Mn\({}^{2+}\) to cell walls. When pH increased the higher removal rate was reached, which could be connected with more negatively charged cells available, promoting a greater metal uptake [34]. Another reason of this observation is that under acidic conditions, the carboxyl groups responsible for the retention of metals are protonated and restrict entry of metallic ions, whereas at a higher pH, these groups are negatively charged, facilitating the binding of the metals ions [43; 44]. Other groups which were identified as functional groups involved in biosorption were hydroxyl, carboxyl, amino, ester, sulfhydryl, carbonyl, and phosphoryl groups [45].