## 4 Discussion  
Given the growing interest in the use of biochar in different fields, this study evaluated how biochar interacts with both pathogens and probiotics and thus how it influences their growth, thus providing evidence supporting its use in animal nutrition. One of the key elements is the sorption capacity, which we evaluated in biochar from vine pruning residue on pathogens and probiotic strains. Regarding pathogen microorganisms, we focused on _Escherichia coli_, especially the F4+ and F18+ strains. Since enterotoxic _Escherichia coli_ (ETEC) characterized by F4 and F18 fimbiae are major causes of postweaning diarrhea in pigs and thus lead to significant economic losses, finding a functional ingredient that can help to prevent the development of pathologies is essential [22]. _Lactobacillus_ sp., especially _L. reuteri_ and _L. plantarum_, were chosen as probiotics since these strains are widely used in pigs and have demonstrated the potential to promote growth, improve feed use, and regulate the immune system in pigs [23]. The production of biochar at relatively low pyrolysis temperatures, such as those applied to vine pruning residues, promotes the development of a predominantly mesoporous structure, along with a rough and spongy surface and an overall amorphous carbon matrix. These morphological features are attributed to the partial preservation of the anatomy of the original plant tissue, particularly xylem elements such as fibers and vessels. These structures thus contribute to a hierarchical porosity, which can increase the adsorption capacity of biochar. The irregular surface texture and tubule-like features derived from xylem cells can increase the surface area available for adsorption. The microbial adsorption of biochar is a complex process governed by many chemical and physical factors. These interactions, which are not only linear, are strongly affected by the environmental conditions and context, thus making case-specific analyses necessary to fully understand their dynamics [24]. Our results revealed that the sorption activity of biochar on _E. coli_ was generally strong, with _E. coli_ F4+ exhibiting a higher percentage of sorption than _E. coli_ F18+. At the maximum dose tested (10 mg/mL), biochar exhibited a percentage adsorption of _E. coli_ F4+ of 69.8% in comparison to 56.1% for _E. coli_ F18+. In addition, biochar has been observed to exhibit an adsorption capacity for _Lactobacillus_ sp. At a concentration of 10 mg/mL, biochar has been found here to demonstrate an adsorption capacity of 38.7% and 36.6% for _L. plantarum_ and _L. reuteri_, respectively. The difference in the adsorption capacity of biochar revealed in this study between _E. coli_ and _Lactobacillus_ spp. could be due to the contrasting properties of these bacteria and the chemistry of the biochar. This depends on different factors, including the surface area,  
Figure 5: Probiotic activity of biochar on _L. plantarum_ (**A**) and _L. reuteri_ (**B**) at doses of 150 and 300 \(\mu\)L/mL for five hours of contact. Different letters indicate significant differences at \(p<0.05\) among different concentrations within the same timepoint. -pore structure, functional groups, and environmental conditions, which jointly influence the sorption mechanism. _E. coli_ cells show a typical size of 0.5-2 \(\upmu\)m, which, compared to that of _Lactobacillus_ cells (approximately 2-4 \(\upmu\)m), are better able to fill the pore size of biochar, thus increasing the selectivity towards this pathogen microorganism and demonstrating a greater adsorption efficacy [25]. In addition to the particle size effect previously discussed, another factor influencing the differential adsorption behavior is the surface chemistry of the biochar. A negative point of zero charge (ZPC) indicates that the biochar surface is predominantly negatively charged under neutral pH conditions, which may partially explain the observed adsorption of _Lactobacillus_ sp.