Nisin is an antimicrobial peptide which belongs to the lantibiotic class of bacteriocins (Class I) and has been employed as a food preservative in over 50 countries (Cotter et al., 2005a). Lantibiotics are characterized by the presence of unusual amino acid residues, including the lanthionines which give these peptides their name (Guder et al., 2000; Chatterjee et al., 2005; Cotter et al., 2005a; Willey and van der Donk, 2007). Lantibiotics, including nisin, can have multiple mechanisms of action facilitated through the binding of lipid II and insertion into bacterial membranes (Brotz et al., 1998; Breukink et al., 1999; Hasper et al., 2006). While Nisin A was first discovered in 1928 in fermenting milk cultures (Rogers and Whittier, 1928), five other naturally occurring variants of Nisin have been described. These are Nisin Z, F and Q, which like Nisin A are produced by Lactococcus lactis, and Nisin U and U2 which are produced by Streptococcus sp. There also exist a number of other more distantly related peptides within the Nisin subgroup of lantibiotics, such as subtilin (Jansen and Hirschmann, 1944), ericin A and ericin S (Stein et al., 2002; Piper et al., 2009a), but which are not regarded as Nisin variants. Nisin Z, first isolated from L. lactis NIZO 22186 from a dairy product, differs from Nisin A by just one amino acid in the final active peptide, His27Asn (Mulders et al., 1991) (Fig. 1). Nisin F is produced by L. lactis F10 isolated from a freshwater catfish in South Africa, and it differs from Nisin A with respect to two amino acid residues, His27Asn (as in Nisin Z) and Ile30Val (de Kwaadsteniet et al., 2008) (Fig. 1). Nisin Q, produced by L. lactis strain 61-14 isolated from a river in Japan, contains both of the substitutions observed in Nisin F as well as two additional variations, Ala15Val and Met21Leu (Fukao et al., 2008) (Fig. 1). Two more distantly related peptides, Nisin U and U2, are produced by Streptococcus uberis 42 (Nisin U) and Streptococcus agalactiae D536 (Nisin U2), and differ from Nisin A by 9 and 10 amino acids respectively (Wirawan et al., 2006) (Fig. 1). These peptides are also three amino acid residues shorter than the other Nisin variants (Rollema et al., 1996) (Fig. 1). In addition to the genes which encode these structural peptides (and their accompanying leaders), Nisin production also requires the presence of genes required for transport (nisT; Nisin A designations are employed), leader cleavage (nisP), post-translational modification (nisB and nisC), regulation (nisRK) and immunity (self-protection; nisI and nisEFG) (Guder et al., 2000). While these genes are present in producers of all variants sequenced (the Nisin F and U2 gene clusters have not been sequenced), they can differ in gene order, as is the case with Nisin U (Wirawan et al., 2006) (Fig. 2). The percentage similarity between the individual biosynthetic, regulatory and immunity proteins produced also varies (Fig. 2), with the Nisin U genes being most distantly related to those associated with Nisin A. In recent years it has been shown that Nisin biosynthetic proteins can be harnessed, both in vivo and in vitro. This technology has facilitated the incorporation of lantibiotic-associated post-translational modifications into a range of unrelated peptides (Kuipers et al., 2006; Kluskens et al., 2009; Kuipers et al., 2009; Majchrzykiewicz et al., 2010; for review see Moll et al., 2010). In addition to the long established use of Nisin as a food preservative, the high potency and multiple mechanisms of action have also been the focus of studies designed around using Nisin in clinical or veterinary applications against drug-resistant pathogens. The efficacy of Nisin A, and to a much lesser extent Nisins Z and F, against methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci (VRE) and (heterogeneous) vancomycin intermediate S. aureus[(h)VISA] has been tested (Severina et al., 1998; Giacometti et al., 2000; Morency et al., 2001; Brumfitt et al., 2002; Kuwano et al., 2005; de Kwaadsteniet et al., 2008; De Kwaadsteniet et al., 2009; Piper et al., 2009b). In this study, we use a bioengineering approach to convert a Nisin A-producing strain into a producer of Nisins F, Q and Z. This bank of bioengineered strains provided us with the opportunity to directly compare the antimicrobial activity of these four natural Nisin variants.