The tail tape measure protein (TMP) of tailed bacteriophages (also called phages) dictates the tail length and facilitates DNA transit to the cell cytoplasm during infection. Here, a thorough mutational analysis of the TMP from lactococcal phage TP901-1 (TMPTP901-1) was undertaken. We generated 56 mutants aimed at defining TMPTP901-1 domains that are essential for tail assembly and successful infection. Through analysis of the derived mutants, we determined that TP901-1 infectivity requires the N-terminal 154 aa residues, the C-terminal 60 residues and the first predicted hydrophobic region of TMPTP901-1 as a minimum. Furthermore, the role of TMPTP901-1 in tail length determination was visualized by electron microscopic imaging of TMP-deletion mutants. The inverse linear correlation between the extent of TMPTP901-1-encoding gene deletions and tail length of the corresponding virion provides an estimate of TMPTP901-1 regions interacting with the connector or involved in initiator complex formation. This study represents the most thorough characterisation of a TMP from a Gram-positive host-infecting phage and provides essential advances to understanding its role in virion assembly, morphology and infection.

Lactococcus lactis is the most widely exploited microorganism in global dairy fermentations. Lactococcal strains are described as typically harboring a number of prophages in their chromosomes. The presence of such prophages may provide both advantages and disadvantages to the carrying host. Here, we describe the deliberate generation of three distinct lysogens of the model lactococcal strain 3107 and the impact of additional prophage carriage on phage-resistance and anti-microbial susceptibility. Lysogen-specific responses were observed, highlighting the unique relationship and impact of each lysogenic phage on its host. Both homologous and heterologous phage-resistance profiles were observed, highlighting the presence of possible prophage-encoded phage-resistance factors. Superinfection exclusion was among the most notable causes of heterologous phage-resistance profiles with resistance observed against members of the Skunavirus, P335, P087, and 949 lactococcal phage groups. Through these analyses, it is now possible to identify phages that may pursue similar DNA injection pathways. The generated lysogenic strains exhibited increased sensitivity to the antimicrobial compounds, nisin and lysozyme, relative to the parent strain, although it is noteworthy that the degree of sensitivity was specific to the individual (pro)phages. Overall, the findings highlight the unique impact of each prophage on a given strain and the requirement for strain-level analysis when considering the implications of lysogeny.