## 4 Discussion  
Beyond technological development, demand is currently raising for the identification of novel lactic acid bacteria (LAB) as potent probiotic candidates [22; 23]. In the present study, LAB was isolated from fermented alfalfa samples, identified by biochemical and molecular methods and characterized in-vitro for recognized probiotic properties including acidity, bile salt tolerance and antimicrobial and anti-oxidant activities are key features to consider bacteria as potent probiotics. In the present study gram-positive, catalase-negative, non-sporulated and rod-shaped bacterial strains were isolated and characterized. The primary result confirmed that these bacteria corresponded to the lactobacteriaceae family. These bacteria were able to ferment the different types of carbohydrates. The carbohydrates fermentation ability of these strains varied among the isolated strains. In addition, isolates secreted different type's extracellular enzymes which are closely associated with proteolytic, glycolytic and lipolytic functions. These enzymes might be useful in nutrient digestibility, which provide additional beneficial characteristic to a probiotic. The extracellular enzyme productions have been varied among the LAB [24]. It might be a reason behind the development of species-specific probiotics. Sensitivity to antimicrobial substances is much important criteria for the selection of probiotic bacteria [25]. All isolates were significantly susceptible to antibiotics such as Chloramphenicol, Nitrofurantoin, Streptomycin, and Dicloxacillin etc. The antibiotic susceptible/ resistance is variable with different LAB, some of the LAB had resistance to ampicillin, vancomycin, chloramphenicol, streptomycin, neomycin, nalidixic acid, gentamycin, kanamycin and novobiocin [26]. In the current study, the isolates exhibited resistance against few antibiotics such as Amikacin Tetracycline and Kanamycin. However, KCC-37 and KCC-38 exhibit high sensitivity against most of the commonly available antibiotics. The secretion of gastric acid in the stomach constitutes a primary defense mechanism. All ingested microorganisms must be overcome these situations, including probiotics [27]. The LAB isolates from alfalfa were screened for their ability to survive in the gastrointestinal tract (GIT) environments. The isolated LAB significantly survived in acidic nature of the stomach and duodenal conditions. Isolated strains showed comparatively lesser survival rates than the standard strain. For the bile salt test, both isolated strains have survived in the bile salts environment of small intestinal conditions. We noted that the KCC-37 survived a higher percentage in bile salt as compared to KCC-38 and standard strain. However, both isolated strains have the ability to cross and survive in GIT environments; it is an essential criterion for the selection of good probiotic bacteria. Cell surface properties by hydrophobicity and autoaggregation are specific parameters for probiotic bacterial adhesion to endothelial cells in the  
Figure 5: Microbial populations, acidification of silages and Organic acids profiles in fermented low moisture IRG silage. (**A**) Microbial population in _L. plantarum_ KCC-37, and _L. plantarum_ KCC-38 inoculated and non-inoculated low moisture silage; (**B**) pH of the low moisture silages (**C**) organic acids including lactic acid (LA), acetic acid (AA), succinic acid (SA) in the experimental silages. Data represent the mean \(\pm\) STD (\(n=3\)), * Significance between treated and untreated group at \(p<0.05\) level. * Significance between the mixed culture treated, single culture treated and control groups at \(p<0.05\) level. intestine and can able to function in the gut environment [28]. In the current study, both _L. plantarum_ showed good hydrophobicity property towards hydrocarbons chloroform and xylene and all strains exhibited significant autoaggregation features.