Parasporins (PS), a class of non-insecticidal and non-hemolytic crystal proteins of Bacillus thuringiensis (Bt), are being explored as promising anti-cancer agents due to their specific toxicity to cancer cells. This work is considered as a first initiative aiming at investigating Algerian soil Bt isolates' activity and cytotoxic potential against cancer cells. A total of 48 Bacillus spp. were isolated from different sites in Algeria. Phenotypic and biochemical tests, 16S rDNA molecular identification, and microscopic observation of crystal have confirmed the identification of Bt for ten strains. A screening for non-hemolytic crystalline proteins was performed. Extraction, purification, and activation of non-hemolytic proteins by chromatographic analysis yielded several polypeptides of different molecular weights. A purified PS1, with pro-protein of 81 kDa and several peptides with different molecular weights (18-58 kDa) after activation by trypsin, has been identified from the strain BDzG. The NH2-terminal sequence deciphered in BLAST analysis showed homology to a Bt PS1 protein. Moreover, the screening of parasporin-1 (PS1) gene has also been performed. Cytocidal activity against human epithelial type 2 (HEp2) cells, considered to originate from a human laryngeal carcinoma, was observed with an IC50 equal to 2.33 μg/ml, while moderate cytotoxicity against adenocarcinomic human alveolar basal epithelial (A549) cells has been shown with IC50 equal to 18.54 μg/ml. No cytotoxicity against normal cells was noted. Fluorescence microscopy revealed a condensed or fragmented chromatin indicating the apoptotic death of HEp2 cells. Thus, Bt PS-producer isolated from Algerian soil might have a potential to join the arsenal of natural anti-cancer drugs with high therapeutic potential.