Recently, the attention to exploring and applying probiotics has been increased. Probiotics are living microscopic organisms that have valuable impacts on health when consumed in adequate amounts. One of the most popular widespread utilized probiotics for many years until now is the Lactobacillus species which exemplifies the hugest heterogeneous group among the lactic acid bacteria (LAB) that normally occupy many environments that contain high nutrient levels such as soil, humans, waters, plants, animals and even air without causing any illness. The purpose of this work is to review and demonstrate the wide diversity of lactobacilli isolation sources and related health benefits applications of the strains discovered in order to break the familiar sources of lactobacillus spp., thereby encouraging more research into unconventional sources and opening up opportunities for clinical applications of Lactobacillus spp. As a result, the isolation sources of lactobacillus spp. in this review have been split into two groups based on the environmental condition, including fermented and non-fermented habitats described as lactobacilli probiotic sources. The unique Lactobacilli isolated from various sources are regarded as potential therapies for a variety of illnesses and physiological abnormalities, including immunomodulation, suppression of carcinogenesis, and cholesterol reduction, in addition to their capacity to guard against infections. This will be highly significant for revealing updated information about Lactobacillus spp. that has attracted considerable attention due to its potential application in clinical practices.