## 3 Results and Discussion  
### Identification and Characterization of Lactic Acid Bacteria  
Lactic acid bacteria were isolated from the queen scallop (_Aequiepecten opercalaris_) digestive tract and the five isolated strains were identified as _Lactiplantiobacillus plantarum_ I, _Lpb. plantarum_ 1, _Lpb. plantarum_ 2, _Lactococcus lactis_, and _Lactiplantiobacillus brevis_, with a high MALDI-TOF score in the range 2-3 indicating a very reliable identification. The strains were also previously tested with the API biochemical method and the results showed a high percentage of identification in the range of 95-99%. The strains were evaluated for their antimicrobial activity against common marine pathogens, and the results demonstrated effective inhibitory activity against all the tested microorganisms (Figure 1). The best antimicrobial activity was detected for the _Lpb. plantarum_ I strain, which inhibited all pathogens in a range of 77-85%. The highest survival in the test microorganisms was detected after treatment with the _Lpb. brevis_ supernatant and was 52-72%. Among all the test microorganisms, bacteria _E. coli_ had the lowest percentage of inhibition with a mean value of 47%. These results are in agreement with previous research, where LAB isolated from marine shellfish showed good antimicrobial activity against marine pathogens [33; 40]. For comparison, in the research by [PERSON] et al. [41] when determining the antimicrobial activity of nine LAB isolates from the digestive tract of _Nile tilapia_ (_Oreochromis niloticus_) against the most common fish pathogens, only one isolated strain, _Lysinibacillus_ sp. 38 HT, successfully inhibited the growth of all pathogens, while the most resistant pathogen proved to be _S. aureus_ inhibited by only three LAB isolates.  
LAB not only exhibit an inhibitory effect through the production of organic acids, such as lactic acid, which leads to a decrease in pH, but they also generate various antimicrobial compounds. These compounds include hydrogen peroxide, CO\({}_{2}\), diacetyl, acetaldehyde, D-isomers of amino acids, reuterin, and bacteriocins [42]. Since the supernatant of tested LAB was neutralized to avoid the inhibitory effect of acid, good activity toward pathogen growth had to be further examined.  
Further testing of the five isolates included the determination of the optimal growth temperature and pH, as well as safety parameters. The optimal temperature for all five strains was 28 \({}^{\circ}\)C with a high number of viable cells ranging from 10\({}^{8}\)-10\({}^{9}\) CFU/mL, which is a minimal reduction compared to the initial number of 10\({}^{10}\) CFU/mL. The Lpb. Brevis, L. lactis, and Lpb. plantarum_ I showed good survival at lower temperatures (\(10^{5}\)-\(10^{7}\) CFU/mL), as well as at a higher temperature of 45 \({}^{\circ}\)C with the number of viable cells ranging from \(10^{4}\)-\(10^{8}\) CFU/mL. The good survival rate of LAB at low and higher temperatures contributes to a much better industrial application of the strains. The optimal pH value for all strains was 6, which is in accordance with the results by [PERSON] and [PERSON] [43], who state that maximum bacterial growth is achieved at pH 5-6 with possible exceptions depending on the strain. Further, LAB isolated from marine animals are usually tolerant to acidic conditions, growing optimally at a neutral or slightly acidic pH [44].  
As far as safety criteria are concerned, all isolates showed sensitivity to the tested antibiotics, as well as the inability to synthesize histidine and tyramine, and the growth test on blood agar showed that they do not have the ability to degrade red blood cells. It is of utmost importance that isolates with future animal or human use are safe for the host, and previous research into the safety aspects of marine LAB supports the results of this work [33; 45; 46].  
Figure 1: Cont.  
Figure 1: _Cont._