Bacteriocins produced by lactic acid bacteria (LAB) are expected to be safe antimicrobial agents. While the best studied LAB bacteriocin, nisin A, is widely utilized as a food preservative, various novel ones are required to control undesirable bacteria more effectively. To discover novel bacteriocins at the early step of the screening process, we developed a rapid screening system that evaluates bacteriocins produced by newly isolated LAB based on their antibacterial spectra and molecular masses. By means of this system, various novel bacteriocins were identified, including a nisin variant, nisin Q, a two-peptide bacteriocin, lactococcin Q, a leaderless bacteriocin, lacticin Q, and a circular bacteriocin, lactocyclicin Q. Moreover, some LAB isolates were found to produce multiple bacteriocins. They were characterized as to their structures, mechanisms of action, and biosynthetic mechanisms. Novel LAB bacteriocins and their biosynthetic mechanisms are expected for applications such as food preservation and peptide engineering.