Lactic acid bacteria, especially Lactobacillus spp., have been considered as excellent probiotic microorganisms, because of their activities in reducing the enteric diseases and maintaining healthy poultry.The current study aimed to evaluate the phenotypic characteristics and the probiotic potentials of Lactobacillus spp. isolated from poultry.A total of 168 lactic acid bacteria (LAB) were isolated from healthy six and twenty-one-day old chickens and their feed samples. The isolated bacteria were identified by morphological, biochemical, and molecular tests including Polymerase Chain Reaction (PCR) and 16S rRNA gene sequencing. Biochemical fingerprinting with Phene Plate system (Ph-P) was done and the acid and bile resistant lactobacilli were subjected to the antibiotic susceptibility test.Amongst all of the examined LAB, 30.3% were resistant to bile and acid. Most of the isolated LAB (57.1%) belonged to the genus Lactobacillus with Lactobacillus brevis (78.1%) as the dominant species followed by L. reuteri (16.6%), L. plantarum (3%), and L. vaginalis (2%). The remaining isolates were identified as Pediococcus spp. (42.9%). The Ph-P cluster analysis of 75 L. brevis and 16 L. reuteri strains showed high phenotypic diversity. Whilst the results of Ph-P typing from L. reuteri strains showed low phenotypic variations especially among the strains sensitive to acid and bile salts.Overall, the results showed that some of the high potential probiotic LAB species existed in Iranian poultry.