The lactococcal plasmid pNP40 mediates insensitivity to phi c2 by an early-acting phage resistance mechanism in addition to the previously identified abortive infection system, AbiF, in the Lactococcus lactis subsp, lactis MG1614 background. A second abortive infection determinant on pNP40, AbiE, does not confer resistance to phi c2. The early-acting mechanism on pNP40 does not prevent phage adsorption nor does it appear to operate by restriction/modification. Phage DNA was not detected in pNP40-containing cells until 30 min following exposure to phi c2 compared with 5 min in a sensitive host; however, electroporation of phage DNA into resistant hosts resulted in the release of phage progeny from a dramatically elevated number of cells compared with conventionally infected hosts. It appears therefore that pNP40 encodes a novel phage resistance mechanism which blocks DNA penetration specifically for phi c2.