Next Article in Journal
Long Non-Coding RNAs and p53 Regulation
Previous Article in Journal
XRCC3 Gene Polymorphism Is Associated with Survival in Japanese Lung Cancer Patients
Article Menu

Export Article

Int. J. Mol. Sci. 2012, 13(12), 16668-16707; doi:10.3390/ijms131216668

Class IIa Bacteriocins: Diversity and New Developments
Yanhua Cui 1, Chao Zhang 1, Yunfeng Wang 2,*, John Shi 3, Lanwei Zhang 1,*, Zhongqing Ding 1, Xiaojun Qu 4 and Hongyu Cui 2
School of Food Science and Engineering, Harbin Institute of Technology, Harbin 150090, China
State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China
Guelph Food Research Center, Agriculture and Agri-Food Canada, Guelph, ON N1G5C9, Canada
Institute of Microbiology, Heilongjiang Academy of Sciences, Harbin 150010, China
Authors to whom correspondence should be addressed; Tel.: +86-451-8593-5058 (Y.W.); Fax: +86-451-5199-7166 (Y.W.); Tel.: +86-451-8628-2901 (L.Z.); Fax: +86-451-8628-2906 (L.Z.).
Received: 10 September 2012; in revised form: 10 October 2012 / Accepted: 12 November 2012 / Published: 6 December 2012


: Class IIa bacteriocins are heat-stable, unmodified peptides with a conserved amino acids sequence YGNGV on their N-terminal domains, and have received much attention due to their generally recognized as safe (GRAS) status, their high biological activity, and their excellent heat stability. They are promising and attractive agents that could function as biopreservatives in the food industry. This review summarizes the new developments in the area of class IIa bacteriocins and aims to provide uptodate information that can be used in designing future research.
class IIa bacteriocin; lactic acid bacteria; diversity; genetic organization; discovery

1. Introduction

Many Gram-positive bacteria, particularly many lactic acid bacteria (LAB) are known to secrete ribosomally-synthesized peptides or proteins that have antimicrobial activity. These compounds (bacteriocins) have been shown to display inhibitory activity against closely related bacteria [1,2]. Four classes of bacteriocins have been defined based on common characteristics, mainly primary structure, molecular weight, mode of action, heat stability and their genetic properties [1,2]. Among these classes, class II, consisting of small peptides that do not contain modified residues, has been divided further into subgroups. Class IIa bacteriocins are characterized by the occurrence of a highly conserved hydrophilic and charged N-terminal region that has a disulphide bond linkage [1,2]. In some bacteriocins, an additional disulphide bond is present. The unambiguous consensus amino acid sequence of class IIa bacteriocins is the “pediocin box” YGNGV (where V can be replaced by L in some cases) [13]. This consensus sequence is included in the conserved N-terminal region YGNGVxCxK/NxxC (where X is any amino acid) [1,2]. Class IIa bacteriocins show their strong inhibitory effect on Listeria sp. as well as other food spoilage and pathogenic bacteria. They have received much attention due to their generally recognized as safe (GRAS) status, their high biological activity, and their heat stability. These compounds show great promise and are attractive candidates for use as biopreservatives in the food industry [47].

2. Diversity of Class IIa Bacteriocins

To date, there are about 50 different kinds of class IIa bacteriocins that have been characterized to the extent that one can with a high degree of certainty determine whether the bacteriocin differs significantly from other bacteriocins (Supplementary Table 1). These bacteriocins have been isolated from a wide variety of LAB, including Lactobacillus sp., Enterococcus sp., Pediococcus sp., Carnobacterium sp., Leuconostoc sp., Streptococcus sp., as well as Weissella sp. [8,9]. They have also been found in the non-LAB Bifidobacterium bifidum[10,11], Bifidobacterium infantis[12], Bacillus coagulans[13] and Listeria innocua[14]. These bacteriocin-producing LAB have been isolated from various environments, including dairy products, fermented sausages, vegetables, and the mammalian gastrointestinal tract.

The class IIa bacteriocins are initially produced as a protein precursor containing an N-terminal leader peptide. This leader peptide is removed by site-specific proteolytic cleavage during export, to yield the mature bacteriocins [2,15]. These mature bacteriocins rang in length from 25 amino acids for mutacin F-59.1 to 58 amino acids for acidocin A. The classification of Gram-positive bacteriocins is complex and several authors have proposed different classifications based on different criteria [13,1618]. The present direction for defining novel classification schemes of Gram-positive bacteriocins tends to take into account the composition, three-dimensional (3D) structure and mode of action of the bacteriocins. Classification of class IIa bacteriocins have been broadly defined first on the basis of their conserved N-terminal region, the “pediocin box,” and then subdivided into 4 subclasses through sequence alignments of the less conserved C-terminal region [3,17,19,20].

The most recent repertoire of class IIa bacteriocins consists of 28 peptides [3]. In this paper, some class IIa bacteriocins were supplemented, including avicin A [21], bavaricin A [22], curvaticin L442 [23], enterocin CRL35 [24], enterocin HF (P86183), bifidocin B [10,11], ubericin A [8], weissellin A [25], bacteriocin 602 [26], bacteriocin 1580 [26], bacteriocin 37 [26], bavaricin MN [27], bacteriocin (P86291.1), bacteriocin E50-52 [28], acidocin A [29], bacteriocin OR-7 [30], bacteriocin L-1077 [31], mundticin L [32], leucocin B [33], prebacterioncin SkgA2, bacteriocin MC4-1 [34], and duracin GL. The 3D structures of bacteriocins were evaluated by SWISS-MODEL Workspace [3537]. The 50 class IIa bacteriocins were classified into eight groups on the basis of their conserved primary structures, 3D structures and mode of action (See Figure 1). The results showed high consistency with the classification of class IIa bacteriocins that were described earlier and discussed by Nissen-Meyer et al.[3] (see Supplementary Table 1).

Group I contains 24 bacteriocins with a sequence length of between 25 and 49 amino acid residues. These peptides are secreted by 17 species of seven genera, including Bacillus sp., Bifidobacterium sp., Carnobacterium sp., Enterococcus sp., Lactobacillus sp., Leuconostoc sp., and Weissella sp. The bacteriocins in this group belong to subgroup 1 which was described in the classification of Nissen-Meyer et al.[3]. The bacteriocins of group I have a flexible hinge at the conserved Asp 17residue. This group can be further subdivided into three subgroups according to their sequence similarities and differences.

Subgroup I-1: includes avicin A, bavaricin A, curvaticin L442, enterocin CRL35, enterocin HF, listeriocin 743A, mundticin, mundticin CRL35, mundticin L, piscicocin CS526, piscicolin 126, sakacin P, and sakacin X. Members of this subgroup exhibit a common consensus motif IGNNxxANxxTGG located at the C-terminal region. Avicin A is produced by Enterococcus avium XA83 which was isolated from feces of healthy infants, and is a probiotic bacterium with diverse antimicrobial potential [21]. Mundticin L is virtually identical to enterocin CRL35. The only difference in sequence occurs in the fifth amino acid residue of the conserved sequence (YGNGX) of these mature bacteriocins, but this change has no influence on antimicrobial activity [32]. Sakacin P is produced by several L. curvatus strains LTH1174, L442 and CRL 705, which were isolated from Greek fermented sausages and fermented meat [38,39]; and by several Lactobacillus sakei strains I151 and LTH673 isolated from sausage and fermented meat [40,41].

Subgroup I-2 encompasses bifidocin B, coagulin, pediocin PA-1, which are produced by B. bifidum, B. coagulans, Enterococcus faecium, Lactobacillus plantarum, Pediococcus acidilactici, Pediococcus pentosaceus and Streptococcus mutans. The common consensus of this subgroup is KYYGNGVTCGK(L)HS(D)CS(R)VDW(R)GKATT(C)C(G)IINNG.

Pediocin PA-1/AcH is a 44-amino-acid class IIa bacteriocin produced primarily by strains of the genus Pediococcus, including Pediococcus acidilactici strains PAC1.0 [42], H [43,44], E, F, M [45,46], K10 [47], HA-6111-2, HA-5692-3 [48], MM33 [49]; Pediococcus parvulus ATO34, ATO77 [50] and P. pentosaceus FBB61 [51]. Pediocin PA-1/AcH is also synthesized by L. plantarum WHE92 [52], L. plantarum DDEN 11007 [53] and E. faecium Acr4.

The genetic determinants for the biosynthesis of pediocin PA-1/AcH are located within a plasmid-borne operon cassette in all producing lactic acid bacterial strains examined to date. In several strains, the sizes and organization of the various pediocin-encoding plasmids are similar [5459]. It has been shown that the plasmids responsible for production in P. acidilactici H can be transferred intragenerically by conjugation [60]. The pediocin PA-1/AcH is the only class IIa bacteriocin for which both cross-species and cross-genera synthesis are known to occur [61].

The entire amino acid sequences of curvaticin L442 and bifidocin B have not been determined and the reported sequence for the bifidocin B contains some uncertainties. The mature sequence of enterocin CRL35 is identical to that of mundticin CRL35, but their leader sequences have some differences. The mature sequence of leucocin A was identical to that of leucocin B and they also had differences in their leader sequences. Sakacin P was identical to bavaricin A, and the peptide we list as sakacin P was a variant of sakacin P.

Coagulin is produced by no-LAB B. coagulans [13]. Interestingly, coagulin is almost identical to pediocin PA-1/AcH, showing 97.7% identity with pediocin PA-1/AcH. More specifically, the coagulin encoding DNA (coaABCD operon) showed 99% identity to that of the papABCD operon encoding the pediocin PA-1/AcH genes [62] (see Figure 2). A putative mob-pre (plasmid recombination enzyme) gene was identified in the coagulin-encoding plasmid pI4[13]. The mob-pre genes present on several plasmids extracted from various Gram-positive genera, including Bacillus, Lactococcus, Streptococcus, Lactobacillus, Enterococcus, and Staphylococcus[13]. In several cases, the corresponding mob genes have been shown to be required for conjugative mobilization and site-specific recombination [63]. Therefore, it was speculated that horizontal gene/operon transfer between P. acidilactici and B. coagulans was possible despite they being relatively unrelated, one is LAB, and the other is no-LAB [13,62].

Interestingly, mutacin F-59.1 from Streptococcus mutans 59.1 shared the conserved sequence KYYGNGVTCGKHSxSVDWxKXT [9]. S. mutans is a human indigenous oral bacterial species. It possesses an advantage against competitive species living in the same niche because of its bacteriocins [64]. The mutacin F-59.1 has a wide activity spectrum inhibiting human and food-borne pathogens [9]. Some amino acids of mutacin F-59.1 have not been determined.

In this subgroup, the bacteriocin-producing strains B. bifidum NCFB 1454 (bifidocin B) and P. acidilactici MM33 (pediocin PA-1), are from human intestinal origin [49,65]. They could be developed for their probiotic properties and as inhibitors of pathogenic bacteria in the gut. Pediocin PA-1 from L. plantarum DDEN 11007 and pediocin A from P. pentosaceus FBB61, are produced by bacteria with established probiotic properties [51,53,66].

Bifidocin B is the first class IIa bacteriocin from a member of the genus Bifidobacterium, sharing 56.8% homology with coagulin and inhibiting the growth of some species of the genera Listeria, Bacillus, Enterococcus, Lactobacillus, Leuconostoc and Pediococcus[11]. Recently, a new bacteriocin bifidin I from Bifidobacterium sp. was reported. Bifidin I from B. infantis BCRC 14602 and showed similarity with bifidocin B, but its whole sequences has not been determined [12]. Bifidin I showed a broad spectrum antimicrobial activity against Gram-positive bacteria and Gram-negative bacteria, including some food-borne pathogens, such as Staphylococcus aureus, Bacillus subtilis, Bacillus cereus, Listeria monocytogenes, Clostridium butyricum, Salmonella enteritidis, Salmonella enterica ssp., and Shigella dysenteriae[12].

Subgroup I-3 is represented by leucocin C, and weissellin A, which are produced by Leuconostoc mesenteroides, Streptococcus uberis and Weissella paramesenteroides. The common consensus of this subgroup is NYGNG(X)2C(X)4CXVXW(X)6IXNNS(X)3GLTG.

Leucocin C and leucocin C-TA33a are produced by different strains of L. mesenteroides, but they showed similar sequences [67]. Leucocin C-TA33a is from L. mesenteroides TA33a, which produced three bacteriocins (leucocin C-TA33a, leucocin B-TA33a and leucocin A-TA33a) with different inhibitory activity spectra [68,69]. The related research revealed that production of leucocin A-, B- and C-type bacteriocins was widespread in Leuconostoc/Weissella strains, including Leuconostoc carnosum LA54a, W. paramesenteroides LA7a, and Leuconostoc gelidum UAL 187-22 [68]. Weissellin A is a unique 4450 Da peptide which is produced by W. paramesenteroides DX which was isolated from a traditional Greek sausage. This bacteriocin exhibits strong activity against L. monocytogenes, Listeria inocua and Clostridium sporogenes[25].

Subgroup I-4 is represented by bacteriocin 602 [26], bavaricin MN [27], divercin V41, divergicin M35, duracin GL, enterocin A, which come from Carnobacterium divergens, Enterococcus durans, E. faecium, L. sakei and Paenibacillus polymyxa. The common consensus of this subgroup is YYGNGV(L)YC.

Group II contains bacteriocin 31, bacteriocin RC714, enterocin SE-K4, bacteriocin T8 (hiracin JM79), penocin A, bacteriocin 1580 and carnobacteriocin B2. The common consensus of this group is YGNGL(V)xCxKxxCxVxW. The bacteriocins in this group belong to subgroup 4 which was described in the classification of Nissen-Meyer et al.[3]. Most class II bacteriocin precursors contain a double-glycine-type signal peptide, and are translocated by dedicated ABC transporters and accessory proteins. However it is likely that some of these bacteriocins contain a different signal peptide. The sequence of hiracin JM79 is identical to that of bacteriocin T8. Hiracin JM79 is produced by Enterococcus hirae DCH5 isolated from wild mallard ducks, and contains a typical sec signal peptide that is believed to direct bacteriocins to the sec translocase embedded in the cytoplasmic membranes [70]. The bacteriocin 31, bacteriocin RC714 and enterocin SE-K4 are also sec-dependent class II bacteriocin [71,72].

Group III contains 10 bacteriocins, which can be further subdivided into two subgroups according to their sequence similarities and differences. The bacteriocins in this group belong to subgroup 2 which was described in the classification of Nissen-Meyer et al.[3].

Subgroup III-1, represented by 8 bacteriocins (bacteriocin MC4-1, leucocin A, leucocin B-Ta11a, mesentericin Y105, plantaricin 423, plantaricin C19, prebacteriocin SkgA2, and sakacin G) has a conserved N-terminal region YYGNGxxCxxxxCxVNWGxA. Plantaricin 423 is bactericidal for many Gram-positive food-borne pathogens and spoilage bacteria, including Listeria spp., Staphylococcus spp., Pediococcus spp., Lactobacillus spp. and so on [73]. Structurally, the N terminus of leucocin A (LeuA) consists of a three-strand antiparallel β-sheet (residues 2–16) that is rigidified by this (9-14)-disulfide moiety [74]. Bacteriocin MC4-1 and prebacteriocin SkgA2 are similar to leucocin A and leucocin A variant (C9L, C14L) in the 3D structures. There structures were determined by the SWISS-MODEL Workspace [3537,75].

Subgroup III-2 consists of lactococcin MMFII and bacteriocin (P86291.1). Lactococcin MMFII is produced by Lactococcus lactis MMFII, which was isolated from a traditional Tunisian cheese [76]. Lactococcin MMFII is the first class IIa bacteriocin produced by a lactococcal strain. It has activity against closely related Gram-positive bacteria, including Lactococcus lactis subsp. cremoris, Lactococcus lactis subsp. lactis, Lactobacillus delbrueckii, Lactobacillus casei, E. faecium, Enterococcus faecalis, and Listeria ivanovi. The bacteriocin (P86291.1) is also produced by Lactococcus sp., showing 90.2% identity with lactococcin MMFII.

Group IV contains carnobacteriocin BM1, curvacin A, enterocin P and ubericin A. This group has the conserved sequences YGNGV(L)YCNxxKCWVNxxE. The group IV bacteriocins lack the hairpin-stabilizing tryptophan and/or cysteine residues that are present at or near the C-terminal end in most class IIa bacteriocins [3]. Carnobacteriocin BM1 is produced by Carnobacterium piscicola LV17B, which is isolated from fresh pork [77]. Curvacin A is produced by Lactobacillus curvatus LTH 1174, which originates from fermented sausage [78]. Enterocin P is produced by several E. faecium strains: IJ-31, P13, GM-1, ATB 197a, JCM5804T, LHICA 51, LHICA 28-4, and LHICA 40-4, which were isolated from various environments, such as fermented sausage, dairy products, feces of newborn infants, and non-fermented animal foods [7984]. Enterocin P showed strong inhibitory action toward Listeria sp. It was processed and secreted by the sec-dependent pathway [79]. Ubericin A is the first streptococcal class IIa bacteriocin to be characterized [8]. It is composed of 49 amino acids with an YGNGL motif at the N-terminal half [8]. Although ubericin A showed high similarity with bacteriocins of subgroup I-3 in amino acid sequences, it showed high similarity with curvacin A in its 3D structure that was determined by SWISS-MODEL Workspace [3537].

The bacteriocin E50-52, bacteriocin 37 and bacteriocin L-1077 are very different and form their own separate group. Bacteriocin E50-52 is produced by E. faecium NRRL B-30746, and shows diverse antimicrobial activity against both Gram-negative and Gram-positive bacteria, including Campylobacter jejuni, Yersinia spp., Salmonella spp., Escherichia coli O157:H7, S. dysenteriae, Morganella morganii, Staphylococcus spp., and Listeria spp. [28]. Bacteriocin 37 is produced by P. polymyxa NRRL B-30507, isolated from broiler chicken, and hasstrong antimicrobial activity against C. jejuni[26]. Bacteriocin L-1077 is produced by Lactobacillus salivarius 1077 (NRRL B-50053), isolated from poultry intestinal materials, and has broad-spectrum antimicrobial activity against 33 bacterial isolates (both Gram-negative and Gram-positive bacteria), including L. monocytogenes A 9-72, E. coli O157:H7, Pseudomonas aeruginosa 508 [31].

The group VII bacteriocins consists of acidocin A and bacteriocin OR-7. This group has a conserved N-terminal region KTYYGTNGVHCTKxSLWGKVRLKN and conserved C-terminal region ILLGWATGAFGKTFH. Acidocin A is produced by L. acidophilus with activity against L. monocytogenes and other closely related Gram-positive bacteria [29]. Bacteriocin OR-7 has 65.5% amino acids sequence similarity with acidocin A with a C-terminal region that is 100% identical to that of acidocin A. Interestingly, bacteriocin OR-7 has different antimicrobial activity from acidocin A. It is active against both Gram-negative and Gram-positive bacteria [30] and has strong antimicrobial activity to Gram-negative bacterium C. jejuni in the chicken gastrointestinal system [30].

The bacteriocin OR-7 and acidocin A have some differences with other class IIa bacteriocins. As a result there is a divergence of opinion as to whether bacteriocin OR-7 and acidocin A should be placed in the class IIa family of bacteriocin [3,19,29,30]. The position of the second cysteine is very different from the very conserved position of this cysteine in the class IIa bacteriocins, suggesting that bacteriocin OR-7 and acidocin A have a different 3D structure in their N-terminal region than the well conserved 3-stranded antiparallel β-sheet like structure which seems to be conserved in most class IIa bacteriocins [3]. Moreover, the sequence and length of the C-terminal region of bacteriocin OR-7 and acidocin A are also very different from other class IIa bacteriocins.

Both bacteriocin OR-7 and acidocin A contained a “pediocin box”-like motif, YGNGVXCXnV, in the N-terminal region of the peptide typical of class IIa bacteriocins, except that a T was present as YGTNGV in the sequence [29,30]. Based on our assessment of previous studies, we are in agreement that bacteriocin OR-7 and acidocin A belong to class IIa family [19,29,30].

3. Biosynthesis of Class IIa Bacteriocins

At least four genes are required for the production of class IIa bacteriocins, including a bacteriocin structural gene encoding a precursor, an immunity gene encoding an immunity protein, genes encoding an ATP-binding cassette transporter and an accessory protein for extracellular translocation of bacteriocin [2].

The class IIa bacteriocin production was regulated by quorum sensing (QS) system. QS systems are present in the majority of Gram-positive and Gram-negative bacteria, as one primary mechanism for bacteria to monitor the environment for other bacteria and to alter behavior on a population-wide scale in response to changes in the number and/or species present in a community [8587].

QS systems used for the regulation of class IIa bacteriocin production are composed of three gene products, including an inducer peptide, a membrane-associated histidine protein kinase (HPK), and a cytoplasmic response regulator (RR) [88]. The inducer peptide is ribosomally synthesized at low levels as a precursor which appears not to be biologically active and contain an N-terminal extension or leader sequence [89]. Subsequent cleavage of the precursor at a specific processing site removes the leader sequence from the antimicrobial molecule concomitantly. Then inducer peptide is secreted and exported through the dedicated transport system involving an ABC-type translocator and an accessory protein [15,88,89]. The presequence of the bacteriocin plays a dual role in bacteriocin biosynthesis [2]. One is a protective role at the cytosolic side of the cell membrane by keeping the bacteriocin inactive. The other is as a recognition signal during export [2].

At a certain concentration threshold of the externalized inducer peptide, the transmembrane HPK detects a change in environmental signal and is activated, leading to its autophosphorylation [88,90]. Then the phosphorylated HPK transfers a phosphate group to its cognate RR. The phosphorylated RR acts as a transcriptional activator and activates expression of bacteriocin-related genes, including genes encoding bacteriocin, immunity protein, secretory apparatus, and regulatory proteins [2,88]. Bacteriocin and immunity genes most often reside on the same operon and are expressed concomitantly. The bacteriocin producer cells protect themselves from their own bacteriocin by the immunity protein. At a certain time, essentially all bacteriocin producer cells in the population are believed to secrete bacteriocins, and this result in a rapid activation of the bacteriocin production [89].

4. Genetic Organization of DNA Coding for Class IIa Bacteriocins

Generally, most class IIa bacteriocin genes are arranged in one or a few operons, which include a bacteriocin structural gene encoding a precursor, an immunity gene encoding an immunity protein, genes encoding an ATP-binding cassette transporter and an accessory protein for extracellular translocation of bacteriocin, and in several cases two regulatory genes encoding a two component system for regulations of the biosynthesis of bacteriocin [19] (Figure 2).

Production of bacteriocins is often correlated with the presence of a plasmid. Several class IIa bacteriocins, for example, enterocin A, divercin V41, sakacin P, carnobacteriocin B2 and carnobacteriocin BM1, have genes that have been shown to be located on chromosome fragments [19,77,9193]. In many bacteriocin-producing bacteria, the bacteriocin structural gene and other related genes were located in one operon. However, genes encoding immunity and secretion functions may not always be linked to structure genes [89,94].

At the present time, all known class IIa bacteriocins are ribosomally synthesized as precursor peptides with an N-terminal leader sequence. The leader sequences of most bacteriocins contain two conserved glycine residues, which may serve as a recognition signal for protein processing and secretion. This double-glycine-type leader sequences were cleaved and removed by ATP-binding cassette (ABC) transporters and their accessory proteins [2]. However, a few class IIa bacteriocins, including bacteriocin 31, enterocin P, enterocin SE-K4, listeriocin 743A, and hiracin JM79 are secreted by the general sec-dependent export system [14,7072,79,95]. These bacteriocins have a hydrophobic N-terminal sec-dependent leader sequence, which directs the secretory protein to the cytoplasmic membrane and is processed by a signal peptidase during translocation across the cytoplasmic membrane. The related genes for production of these bacteriocins are unknown [14,71,72,79,9598].

Class IIa bacteriocins show a remarkable conservation of gene arrangement (Figure 2). The genetic organization of leucocin A gene cluster (lca locus) from L. gelidum UAL187 is a typical bacteriocin locus [99]. The lca locus includes two different directions operons with four bacteriocin-related genes lcaA, lcaB, lcaC and lcaD. The immunity protein gene lcaB is located immediately downstream of the structural leucocin A gene lcaA. The accessory transporter gene lcaD occurs also downstream of gene lcaC encoding an ABC transporter [99].

The genetic organization of sakacin P gene cluster (spp locus) from L. sakei LTH673 and LTH674 is complicate, when compared to leuconcin A [40,93]. It is composed of three operons, which encode a 61-amino-acid sakacin P precursor SppA, a sakacin P immunity protein SpiA; a transport and secretory system (a 718-amino-acid ABC transporter protein SppT and an accessory factor for ABC transporter protein SppE); and a three-component regulatory system (inducing peptide preprotein SppIP, HPK SppK and RR SppR), respectively [40,93]. The production of sakacin P in L. sakei Lb674 and LTH673 is regulated by a typical peptide pheromone-based QS mechanism [40,93].

The genetic organization of divercin V41 presents an unusual organization [92]. The dvn locus encodes a 66-amino-acid divercin V41 precursor, an ATP dependent transporter, two immunity-like proteins and two components of a lantibiotic-type signal-transducing system [92] (see Figure 2). Interestingly, a so-called transport accessory protein was absent from the locus. Generally, the genes encoding the HPK are located upstream of the genes encoding RR in anti-listeria bacteriocin operon [100]. However, in the dvn locus of divercin V41, the HPK gene followed the RR gene, which is a characteristic of lantibiotic operons. The genetic organization of the fragment suggests important gene rearrangements [92].

Sometimes one locus can include productions of two bacteriocins. L. sakei 5 produces a plasmid-encoded bacteriocin sakacin P, as well as two chromosomally encoded bacteriocins, i.e., sakacin T, which is a class IIb two-peptide bacteriocin and sakacin X, which is a class IIa bacteriocin [101]. The sakacin TX locus encodes structural genes of sakacin T and sakacin X, including two adjacent but divergently oriented gene clusters (See Figure 2). The first gene cluster stxPRKT is believed to encode an inducing peptide, three proteins involved in regulation and secretion of these bacteriocins. The second gene cluster includes sakTα, sakTβ, sakIT, sakX and sakIX, which encode the structural and immunity genes for sakacin T and sakacin X [101].

L. mesenteroides FR52 produces both mesentericin 52A and 52B [102]. Mesentericin 52A is a 37-amino-acid class IIa bacteriocin, identical to mesentericin Y105 from L. mesenteroides Y105 [103]. Mesentericin 52B is a 32-amino-acid atypical class II bacteriocin, identical to mesentericin B105 from L. mesenteroides Y105 [104]. The mes locus of L. mesenteroides FR52 is involved in productions of mesentericin 52A and 52B [104]. The previous study revealed that ATP dependent transporter MesD and transport accessory protein MesE were involved in secretion and transport of these bacteriocins [104]. Mesentericin 52A and mesentericin 52B have own immunity genes mesI and mesH, respectively.

The sakacin G gene cluster (skg locus) from L. sake 2512, R1333 and CWBI-B1365 was very interesting because it contained duplicated structural genes skgA1 and skgA2[105107]. There is only a two-amino-acid difference in sequence occurs in leader peptides of these prebacteriocins which makes these mature peptides, SkgA1 and SkgA2, essentially identical [106,107].

The genetic organization of avicin A gene cluster (avc locus) from E. avium has been established [21]. It is the first bacteriocin locus identified in E. avium to be characterized at the molecular level [21]. The locus showed a particular gene organization. The accessory gene avcD associated with bacteriocin transport did not occur immediately downstream of the gene avcT (which encodes an ABC transporter), but two regulatory genes avcK (which encodes a HPK) and avcR (which encodes a RR) followed the gene avcT[21]. The avcK, avcR, and induction peptide pheromone-encoding gene avcF, constituted a three-component regulatory system in the avicin locus. This indicated that the production of avicin A was regulated by the peptide pheromone-inducible regulatory system [21]. For most class IIa bacteriocins, three genes responsible for regulation are located in the same operon, but avcK, avcR, and avcF were located in two different operons (See Figure 2). In this locus includes two bacteriocins structural genes avcA and avcB. Avicin B is a divergincin-like bacteriocin, but it didn’t show antimicrobial activity and is probably a relic of a previous functional bacteriocin [21].

5. Structure-Function Relationship and Target Recognition of Class IIa Bacteriocins

To date, the 3D structures of leucocin A [74], carnobacteriocin B2 [108], sakacin P [109] and curvacin A [110] have been characterized by nuclear magnetic resonance (NMR) spectroscopy. The 3D analysis revealed that class IIa bacteriocins consist of a hydrophilic, cationic and highly conserved N-terminal β-sheet domain, and a flexible, diverse hydrophobic/amphiphilic C-terminal domain [3,74,108110]. The former is structurally stabilized by a conserved disulfide bridge; the latter contains a central amphiphilic α-helix, ending with a structurally extended C-terminal tail. The amphipathic α-helix was critical for antimicrobial specificity and temperature-dependent activity of these class IIa bacteriocins [74,108,111114]. The C-terminal part of some class IIa bacteriocins, such as enterocin A, divergicin M35, divercin V41, coagulin, pediocin PA-1, sakacin G and plantaricin 423, formed a hairpin structure which was stabilized by a disulfide bridge between a cysteine residue in the middle of the α-helix and a cysteine residue at the C-terminus [3].

Two cysteines that come from the conserved N-terminal region (YGNGVxCxK/NxxC) of class IIa bacteriocins formed a conserved disulfide bond. In most class IIa bacteriocins, the disulfide bond is formed between cysteine9 and cysteine14. Extensive studies indicate that this conserved disulfide bond is required for antimicrobial activity for class IIa bacteriocins [115117]. Mutants of mesentericin Y105 (cysteine9→serine9, cysteine14→serine14) showed a marked loss in antimicrobial effects [115]. The antimicrobial activity of pediocin PA-1 was abrogated by the substitution of 11 different amino acids at cysteine14 based on NNK scanning [116]. Substitution of the cysteines with serines in leucocin A (LeuA) abolished antimicrobial effects [117].

However, some results from Derksen et al. indicated that the disulfide bond in leucocin A (LeuA) could be replaced by a noncyclic diallyl moiety without significant loss in activity [117]. The leucocin A (C9F, C14F), bis-allyglycine-leucocin A, and norvaline-leucocin A retained activities comparable to that of the natural leucocin A [75,114]. The researchers speculated that hydrophobic or π-stacking interactions can compensate for the absence of the disulfide in this molecule and assist receptor binding [75,114,117].

Three analogues of leucocin A (LeuA) and six analogues of pediocin PA-1(Ped) were synthesized by replacing the conserved cysteines that form a disulfide bond with pairs of hydrophobic amino acids [114]. Noncovalent hydrophobic interactions in all of the leucocin A (LeuA) derivatives effectively replaced the disulfide and afforded peptides with full antimicrobial activity [114]. Apparently the propensity of the intraloop sequence of leucocin A (LeuA) to induce β-turns in combination with the hydrophobic interaction of the two Phe residues is sufficient to achieve the appropriate conformation for bioactivity [114,118].

Sit et al. presented the 3D solution structures of the inactive (C9S, C14S)-leucocin A and the active (C9L, C14L)-leucocin A peptides [75]. Mutation of the two cysteine residues to serines or leucines did not affect the overall charge of the peptide, and therefore is highly unlikely to interfere with the electrostatic interactionsbetween the peptide and the bacterial cell surfaces. It was speculated that the N terminus may be serving a more crucial function, such as forming intermolecular contacts with other leucocin A–EIItman complexes during pore formation [75].

Receptor binding might occur on the surface of a three-strand antiparallel β-sheet at the N terminus of the peptide as well as by recognition of the hydrophobic face of the amphipathic C-terminal α-helix, which is known to be required and determines specificity for particular organisms [112,119,120]. These results indicate that although the N-terminal loop has a vital influence on the activity of the peptide, additional interactions at the C terminus with the receptor must match and contribute to the overall activity [115,119121].

Most class IIa bacteriocins present a single intramolecular disulfide bond between cysteine9 and cysteine14. The C-terminal part of a few class IIa bacteriocins, contains an additional C-terminal disulfide bridge, such as sakacin G (between cysteine24 and cysteine37), plantaricin 423 (between cysteine24 and cysteine37), pediocin PA-1/AcH (between cysteine24 and cysteine44), divercin V41 (between cysteine25 and cysteine43), and enterocin A (between cysteine29 and cysteine47). The second disulfide bridge not only plays an important role in stabilizing the 3D structure of the C-terminal domain, but also correlates strongly with spectrum of activity [2,20,109,113,122,123]. The previous studies indicated that the second disulfide bridge in the class IIabacteriocins contributes to widening of the antimicrobial spectrum as well as to higher potency at elevated temperatures [113].

It is well known that class IIa bacteriocins kill target cells by forming pores and disrupting the integrity of target cell membranes, causing dissipation of proton motive force, depletion of interacellular ATP and leakage of amino acids and ions [2,19]. Numerous mode-of–action studies have demonstrated that the sugar transporter mannose phosphotransferase system (Man-PTS) serve as target receptors for class IIa bacteriocins on sensitive cells [124131]. The Man-PTS, which is a complex sugar uptake system in the Gram-positive Firmicutes and Gram-negative Gammaproteobacteria, includes a general PTS protein enzyme I (EI), a histidine containing phosphocarrier protein (HPr) and a carbohydrate-specific protein complex (enzyme II, EII) [132].

The enzyme II consists of four subunits: IIA, IIB, IIC and IID [132]. Subunits IIA and IIB are located in the cytoplasm and are responsible for phosphorylation. They are often found together on one protein. The IIC subunit is an integral membrane protein involved in sugar transport. The IID subunit is also a transmembrane protein [132]. The membrane proteins IIC and IID together form a membrane-located complex. IIA and IIB are in reversible contact with the membrane-located complex [129,133]. Other studies indicated that a single extracellular loop of the membrane-located protein IIC (MptC) was involved in specific target recognition by the class IIa bacteriocins, and was the major determinant responsible for species-specificity [125,130].

The proposed mechanism of action for IIa bacteriocins is as follows: first, the N-terminal β-sheet domain of bacteriocin binds to the extracellular loop of IIC in the Man-PTS. Then, C-terminal α-helix-containing hairpin or hairpin-like domain of the bacteriocin interacts with the transmembrane helices of the Man-PTS, leading to conformational changes in the Man-PTS proteins in a manner that renders the transporter irreversibly open thereby causing uncontrolled efflux of essential molecules, disruption of the membrane integrity and in effect, cell death [131,134]. In bacteriocin producing cells, a cognate immunity protein tightly binds the receptor in a bacteriocin-dependent manner, to prevent killing by the bacteriocin [129]. However some class IIa bacteriocins, including enterocin P and sakacin A, showed a different mode of receptor recognition. They employ the IIC and IID complex as a receptor on target cells and then the cognate immunity protein (LciA) is tightly associated with the bacteriocin-receptor complex to render producer cells immune [129,135].

Most class IIa bacteriocins have a relatively narrow inhibitory spectrum, inhibiting predominantly genera or species closely related to the bacteriocin producers. In order to reveal the mechanism of the receptor function specificity, a phylogenetic analysis of membrane-located proteins (IIC and IID) of 86 Man-PTSs from a wide range of bacterial genera was performed [136]. These man-PTSs are clustered into three distinct groups, named groups I, II and III. Fourteen man-PTSs distributed all over the phylogenetic tree were selected for heterologous expression in L. lactis indigenous man-PTS-deletion mutant [136]. Bacteriocin sensitivity of the different L. lactis clones was determined with four class IIa bacteriocins, including pediocin PA-1, enterocin P, sakacin P, and penocin A [136]. The results indicated that only members of group I could serve as receptors for class IIa bacteriocins. A multiple sequence alignment analysis of IIC and IID proteins revealed three sequence regions (two in IIC and one in IID) that distinguish members of the group from those of the other groups, suggesting that these amino acid regions confer the specific bacteriocin receptor function [136].

The receptor efficiencies of Listeria, Enterococcus, Lactobacillus, Leuconostoc, Carnobacterium, Clostridium, Pediococcus and Streptococcus varied in a pattern directly related to their phylogenetic position [136]. The species of Enterococcus, Listeria and Carnobacterium showed most active receptors and were highly sensitive to four IIa bacteriocins; the species of Lactobacillus, Pediococcus and Clostridium are also frequently inhibited by these bacteriocins, although they are often less sensitive; and the strains of Streptococcus and Leuconostoc are occasionally reported to be sensitive to class IIa bacteriocins at a low level. These results are in line with previous comparative analyses of the inhibitory spectra of class IIa bacteriocins [122,137]. Different strains of the same bacterial species can vary greatly in sensitivity to a given bacteriocin [122,138]. The variation in sensitivity might be due to differential expression levels of the receptor [136].

Generally, the conserved N-terminal region of class IIa bacteriocin was speculated to be involved in the receptor interaction, and the diverse C-terminal region was responsible for target cell species-specificity [136]. But some studies strongly suggest that the C-terminal region of class IIa bacteriocin might be involved in interaction between bacteriocin and its receptor [119,121,139,140]. Therefore it was speculated that N-terminal and C-terminal regions take part in the interaction with target cell receptor and that, they have different function during different stage of interaction. Synthesis of bacteriocin mutants and analogues provides valuable structure-activity relationships and tools to obtain further information on the peptide-receptor complex [117,119].

Resistance of Listeria spp. and other Gram-positive bacteria to class IIa bacteriocins was correlated with loss or reduction of expression of Man-PTS, inthe following phenotypes [132,135,141143]: (i) absence of the IIAB subunit of Man-PTS in the proteomes of resistant bacteria [125,143]; (ii) mutations in the sigma transcription factor σ54 (rpoN) and the σ54-dependent transcription activator ManR of the mpt operon [124,126,127,144146], (iii) a mutation in the promoter proximal mptA (IIA) cistron [125], and (iv) in-frame deletions in the mptD (IID) gene (which may have compromised the folding and stability of IID and IIC) [144]. Recently natural food isolates of L. monocytogenes with different susceptibilities to class IIa bacteriocins were investigated [135]. The results also identified Man-PTS as a key player in the mechanisms of resistance. At the same time, downregulation of the mpoABCD (mannose permease one) operon in L. monocytogenes was shown to promote resistance to class IIa bacteriocins [147]. The mpoABCD operon putatively encodes a PTS permease of the mannose family similar to that encoded by the mpt operon. In silico analysis indicated that mpo transcription might be dependent on σ54.

Bacterial strains sensitive to class IIa bacteriocins readily give rise to resistant mutants upon bacteriocin exposure. The development of highly tolerant and/or resistant strains may decrease the efficiency of bacteriocins as biopreservatives. The acquiring of resistance to bacteriocins can significantly affect physiological activity profile of bacteria, alter cell-envelope lipid composition, and also modify the antibiotic susceptibility/resistance profile of bacteria [148].

6. Discovery of Class IIa Bacteriocins

To date, traditional screening strategies have relied on detection of antimicrobial activity as the basis for discovery of new and potent bacteriocins [131]. New bacteriocins are detected and identified by screening large number of potential bacteriocin-producing bacteria for antimicrobial activity. The screened bacteriocins are then purified and characterized. These classic screening strategies are time-consuming and labor-intensive, so researchers need to explore and develop more rapid and higher-throughput approaches for identification of bacteriocins potential [149152]. The PCR assays that target bacteriocin-coding genes or bacteriocin regulation-related genes for rapid detection of bacteriocins have been developed [152156]. Most PCR assays can only detect known bacteriocins because they use specific primers which were designed according to previously characterized bacteriocins [154,155,157]. Więckowicz et al. have developed a rapid PCR assay with primers which were designed on the basis of a large scale alignment of class IIa bacteriocin genes. Several potentially novel bacteriocin-coding sequences were found by means of this high-throughput PCR assay [152].

A large number of LAB genomes have been published during the last decade [158,159]. At the same time, bioinformatics as well as new technologies such as transcriptomics, proteomics and metabolomic analysis have expanded tremendously in past decade. All of the above mentioned technologies have provided a basis for detection of bacteriocins by means of silico analysis [160]. Recently, there has been a trend from classical screening strategies for antimicrobial activity towards silico analysis of genomic data as computational approaches are able toaccelerate the process of novel antimicrobial peptides (AMPs) discovery and design [131,137,161,162].

Dirix et al. identified over 50 bacteriocins or bacteriocin-like peptides by screening for peptides containing a double-glycine leader sequence and the corresponding ABC transports in 165 fully sequenced bacterial genomes (including 45 Gram-positive bacteria and 120 Gram-negative bacteria) [161,162]. Diep et al. identified a new class IIa bacteriocin penocin A in the genome of P. pentosaceus ATCC 25745 by means of silico-based analysis. The antimicrobial activity of penocin A has been determined by experiments [137]. The silico analysis for prediction of bacteriocins, is a challenging task due to the small sizes and diversity in sequence, structure and function of bacteriocins [131].

Some databases and bioinformatics tools have been developed and designed for prediction of AMPs production by both Gram-positive and Gram-negative bacteria. For example, an antimicrobial peptide database (APD) was developed by means of sequence similarity and certain known principles of AMPs [163]. The database was updated in 2009 [164]. AMPer database provided hidden Markov models (HMMs) to automatically discover AMPs [165]. An integrated open-access database BACTIBASE ( [166], and a genome mining software BAGEL2 ( [167] were specifically designed for AMPs discovery [168,169]. Wang et al. constructed a new method by means of sequence alignment and feature selection methods to predict AMPs [170]. Recently Fernandes et al. employed adaptive neuro-Fuzzy inference system (ANFIS) as a pattern recognition tool to classify a putative peptide as an AMP or non-AMP [171].

Quantitative structure–activity relationship (QSAR) modeling is one of the most broadly used chemoinformatics approaches. It can be defined as quantitative models that correlate the variation in measured biological activity with the variation in molecular structure among a series of chemical compounds. QSAR has been applied successfully to AMPs discovery [172175]. The CAMEL database employed QSAR and artificial neural networks (ANN) to predict AMPs function [176]. Recently a novel quantitative prediction method of AMP was established by QSAR modeling based on the physicochemical properties of amino acids [177].

The activity of an AMP is commonly expressed as the threshold concentration (minimum inhibitory concentration, MIC) upon which bacterial growth is inhibited. Biophysical studies with model phospholipid membranes often identify concentration thresholds upon which the peptide behavior becomes disruptive through pore formation or membrane lysis [178183]. The connections between in vivo MICs and thresholds in model membranes have been recently proposed [183,184]. Recently, Melo et al. developed an interaction model of antimicrobial peptides with biological membranes [178]. A straightforward and robust method was presented and used to implement this relationship. The methodology provides a basis for fast, cost-effective alternatives for screening AMPs, with potential application to high-throughput screening approaches. These tools will accelerate and optimize the discovery and identification of novel bacteriocins. Howerverthese bacteriocins still have to be verified by measuring their antimicrobial activities according to excepted experimental procedures.

7. Conclusions

A large number of new class IIa bacteriocins have been detected and purified in the last decade. Some class IIa bacteriocins with wide-spectrum antimicrobial activity have been reported and new discovery methods have been introduced. Acuña et al. presented a novel procedure for designing hybrid bacteriocins through fusion of microcins with class IIa bacteriocins in order to produce new wide-spectrum bacteriocins with high specific activity [185]. All of these advancements will accelerate the developments of class IIa bacteriocins.

Supplementary Information

Table S1. Some characteristics of the class IIa bacteriocins.
Table S1. Some characteristics of the class IIa bacteriocins.
BacteriocinAccount NucleotideAccount ProteinPrepeptie size (aa)MP size (aa)MP Mass (Da)pIProducerOriginReferences
Group I
Sub-group I-1
Avicin AFJ851402.1ACZ36002.161434291.99.32E. avium XA83Feces of healthy infants[ 21]
Bavaricin A/SppAAF526262AAM88858.161434435.98.76L. sakei MI401Sourdough[ 22]
Curvaticin L442#P84886.1L. curvatus L442Greek fermented sausage[ 23]
Enterocin CRL35AY398693AAQ95741.1584342879.82E. mundtii CRL35Argentinian artisanal cheese[24]
Enterocin HFP861834343339.37E. faecium HS and TA29Humans and fish
Listeriocin 743AAF330821.1AAK19401.1714344849.98L. innocua 743Food[186, 4]
MundticinP80925.1434287E. mundtii ATO6Fresh chicory endive[187]
Mundticin CRL35AY444743AAR26473.1584342879.82E. mundtii CRL35/AT06Artisanal cheese[ 24]
Mundticin KSAB066267BAB88211.1504342879.82E. mundtii NFRI 7393/AT06Fresh chicory endive[188]
Mundticin LFJ899708.1ACQ77507.158434301.89.82E. mundtii CUGF08Alfalfa sprouts[ 32]
Mundticin QU243 *4287E. mundtii QU 2Fermented soybean[189]
Pediocin ACCEL#P. pentosaceus ACCEL
Piscicocin CS526 #C. piscicola CS526Cold-smoked salmon[190]
Piscicolin 126AY812745AAX21354.1624444179.32C.maltaromaticum UAL26Vacuum-packaged beef[191]
Piscicolin 126AF275938.1AAK69419.1624444179.32C. piscicola JG126Spoiled ham[ 192]
Piscicocin V1a4444179.32C. piscicola V1Fish[193]
Sakacin PDQ019413.1AAY44078.161434461.98.74L.curvatus LTH1174Meat fermentation[38]
Sakacin PDQ019414.1AAY44080.161434461.98.74L.curvatus L442Greek fermented sausage[ 39]
Sakacin PAY875983AAW79057.161434435.98.76L.sakei I151Sausage[41]
Sakacin PAF002276.1AAB93970.161434435.98.76L.sakei LTH673Meat fermentation[40]
Sakacin PNZ_AGBU01000084.1ZP_09041901.161434435.98.76L. curvatus CRL 705Fermented sausage
Sakacin XAY206863AAP44569.1614343649.32L. sakei 5Malted barley[101]
Sakacin XZP_09041912.1614343649.32L. curvatus CRL 705Fermented sausage
Sub-group I-2
Bifidocin B #363801.58.05B. bifidum NCFB 1454Human isolate[10,11]
CoaA/Coagulin/CoaAAF300457.1AAG28763.162444614.28.66B. coagulans I4Cattle feces[194,13]
Mutacin F-59.1P86386.125 *S. mutans 59.1[ 9]
PapANC_004832.1NP_857602.162444627.28.66P. acidilactici H[195]
PediocinEU826148.1ACF32966.162444627.28.66P. acidilactici MTCC 5101
Pediocin A4446288.66P. pentosaceus FBB61Cucumber fementations[ 51]
Pediocin AcHS74PEDACHAAA98337.162444627.28.66P. acidilactici HFermented sausage[44]
Pediocin AcH444627.28.66L. plantarum WHE92Soft cheese in France[52]
Pediocin PA-1HQ876214.1AEH68223.162444627.28.66E. faecium Acr4
Pediocin PA-1AAB23877.144 *P. acidilactici[196]
Pediocin PA-1M83924.1AAA25559.162444627.28.66P. acidilactici PAC1.0.Sorghum beer[ 197, 42]
Pediocin PA-14446288.66L. plantarum DDEN 11007[53,66]
Pediocin PA-14446288.66P. acidilactici MM33Human stool[49]
Pediocin PP-1444602.28.66P. pentosaceus CBT8Kimchi[198]
Pediocin SJ-1P. acidilactici SJ-1Meat[ 57]
Prepediocin AcHS44537.1AAC60413.262444605.28.33P. acidila I ctici Lb42-923[44]
Prepediocin PA-1AY705375.1AAT95422.162444627.28.66P. acidilactici K10Kimchi[47]
Leucocin CLCCC_LEUMEP81053.24345958.76L. mesenteroides 6Malted barley[67]
Leucocin C-TA33a36 *4598L. mesenteroides TA33aVacuum-packaged meat[ 69]
Weissellin A4344509.32W. paramesenteroides DX[25]
Bacteriocin 602P86393.13938647.2P. polymyxa NRRLB-30509Broiler chicken, crop[ 26]
Bavaricin MNP80493.242476910.0L. sakei MNMeat[27]
Divercin V41AJ224003CAA11804.166434512.38.65C. divergens V41Fish viscera[92,199]
Divergicin M35P84962.1434518.758.3C. divergens M35Smoked salmon[200]
Duracin GLHQ696461.1ADW93772.171434966.78.74E. durans 41DCheese product
Enterocin AX94181.1CAA63890.1654748298.98E. faecium CTC492Fermented sausage[ 91]
Enterocin A654748338.98E. faecium WHE 81Cheese[ 201]
Enterocin ANZ_GG692545.1ZP_05660016.165474831.68.98E. faecium 1,230,933
Enterocin AAB038464.1BAA92138.165474831.68.98E. faecium N15Japanese rice-bran paste[153]
Enterocin A/ EntAAF099088.1AAD2913265474831.68.98E. faecium DPC1146[202]
Enterocin BC25AF240561.1AAF44686.165474831.68.98E. faecium BC25[203]
Group II
Bacteriocin 31 /BacAD78257.1BAA11329.167435007.89.72E. faecalis YI717Clinical sample[ 72]
Bacteriocin 1580P86394.13534867.8B. circulans NRRLB-30644Broiler chicken, crop[ 26]
Carnobacteriocin B2L47121.1AAB81310.166484969.99.97C. piscicola LV17BPork[77,108]
Bacteriocin 43AB178871BAF36626.174445092.99.26E. faecium[204]
Bacteriocin RC714434936.78.74E. faecium RC714Human fecal[ 205]
Bacteriocin T874445092.99.26E. faecium T8Children Infected with HIV[ 206]
Enterocin SE-K4AB092692.1BAC20326.176485356.29.93E. faecalis K-4Grass silage in Thailand[207,71]
Hiracin JM79DQ664500ABG47453.174445092.99.26E. hirae DCH5Mallard ducks[70]
Penocin A/PenAYP_80363560424688.49.72P. pentosaceus ATCC 25745[137]
Group III
Sub-group III-1
Bacteriocin MC4-1EU047916ABW08100.171434890.69.27E. faecalis MC4[34]
Carnocin CP52CPU76763AAB18989.166484969.99.97C. piscicola CP52Cheese[ 208]
Leucocin AM64371.1/LEULAIPAAA68003.161373932.38.78L. gelidum UAL 187Vacuum-packaged meat[209,33]
Leucocin B-Ta11aS72922.1AAC60488.161373931.68.78L. carnosum Ta11aVacuum-packaged meat[ 33]
Mesentericin 52AAY286003AAP37395.161373869.58.78L. mesenteroides subsp. mesenteroides FR52Raw milk[ 102]
Mesentericin Y105X81803.1CAA57405.161373869.58.78L.mesenteroides Y105Goat's milk in France[103]
Plantaricin 423AF304384AAL09346.156373934.68.67L. plantarum 423Sorghum beer[73, 210212]
Plantaricin C19363845.39.88L. plantarum C19Fermented cucumbers[213, 214]
Prebacteriocin SkgA2ZP_08080540.156384159.89.03L. ruminis ATCC 25644Human gastrointestinal tract
Sakacin GAF395533.1AAM73712.155373837.47.96L. sakei 2512Rhodia food collection[105]
Sakacin GFJ621568.1ACM68469.155373837.47.96L. sakei R1333Smoked salmon[107]
Sakacin GEU570253ACB72724.155373837.47.96L. sakei CWBI-B1365Raw poultry meat[106]
Sakacin GEU570253ACB72725.155373837.47.96L. sakei CWBI-B1365Raw poultry meat[106]
Sub-group III-2
Lactococcin MMFIIP83002.1374144.67.25L. lactis MMFIITunisian cheese[76]
BacteriocinP86291.1414601.37.25Lactococcus sp.
Group IV
Carnobacteriocin BM1L29058.1AAA23014.161434524.68.76C. piscicola LV17BFresh pork[ 77]
Curvacin AS67323.1AAB28845.159414308.09.37L.curvatus LTH 1174Fermented sausage[ 78]
Ubericin AEF203953.1ABQ23939.170495270.59.35S. uberis E[8]
Enterocin PGQ369522.1ACU28817.171444701.37.25E. faecium IJ-31Dairy products in Islamabad[84]
Enterocin PAF005726AAC45870714444938.22E. faecium P13Spanish fermented sausage[79]
Enterocin PAY728265AAU29394.1444714.35.51E. faecium GM-1Feces of a newborn infant[81]
Enterocin P-likeAY633748AAT58220.1444701.37.25E. faecium ATB 197a
Enterocin P-likeAB075741BAC00780.140*E. faecium JCM5804T[ 80]
Enterocin PDQ867125ABI29857.1444629.38.22E. faecium LHICA 51Nonfermented animal foods[82]
Enterocin PDQ867124ABI29856.1444629.38.22E. faecium LHICA 28-4Nonfermented animal foods[82]
Enterocin PFJ416487ACJ46053.1444629.38.22E. faecium LHICA 40-4Nonfermented animal foods[83]
Piscicocin V1b4345268.76C. piscicola V1Fish[193]
Sakacin AZ46867CAA86942.159414308.09.37L. sakei Lb706Meat[215217]
Group V
Bacteriocin E50-52P85148.1394124.98.12E. faecium NRRL B-30746[28]
Group VI
Bacteriocin L-10773734549.1L. salivarius 1077Healthy broiler chickens[31]
Group VII
Bacteriocin 37P86395.1303465.410.1P. polymyxa NRRL B-30507Broiler chicken, crop[ 26]
Group VIII
Acidocin ABAA0712081586501.510.93L. acidophilus TK9201[29]
Bacteriocin OR-754621410.32L. salivarius NRRL B-30514Cecal contents of chickens[30]

aa, Amino acids; MP, Mature peptide;#, the whole sequence of bacteriocin has not been determined, including Curvaticin L442 and bifidocin B;*, some amino acids of bacteriocin has not been determined;B. circulans, Bacillus circulans; B. coagulans, Bacillus coagulans; B.bifidum, Bifidobacterium bifidum; C. divergens, Carnobacterium divergens; C. maltaromaticum, Carnobacterium maltaromaticum; C. piscicola, Carnobacterium piscicola; E. avium, Enterococcus avium; E. durans, Enterococcus durans; E. faecalis, Enterococcus faecalis; E. faecium, Enterococcus faecium; E. hirae, Enterococcus hirae; E. mundtii, Enterococcus mundtii; L. acidophilus, Lactobacillus acidophilus; L. carnosum, Leuconostoc carnosum; L. curvatus, Lactobacillus curvatus; L. gelidum, Leuconostoc gelidum; L. innocua, Listeria innocua; L. lactis, Lactococcus lactis; L. mesenteroides, Leuconostoc mesenteroides; L. pentosus, Lactobacillus pentosus; L. plantarum, Lactobacillus plantarum; L. ruminis, Lactobacillus ruminis; L. sakei, Lactobacillus sakei; L. salivarius, Lactobacillus salivarius; P. acidilactici, Pediococcus acidilactici; P. parvulus, Pediococcus parvulus; P. pentosaceus, Pediococcus pentosaceus; P. polymyxa, Paenibacillus polymyxa; S. mutans, Streptococcus mutans; S. uberis, Streptococcus uberis; W. paramesenteroides, Weissella paramesenteroides; HIV, Human Immunodeficiency Virus.


This work was supported by a grant from the State Key Laboratory of Veterinary Biotechnology (No. SKLVBF201202), and a project in the Postdoctoral Science-Research Foundation of Heilongjiang province (LBH-Q11119).


  1. Klaenhammer, T.R. Genetics of bacteriocins produced by lactic bacteria. FEMS Microbiol. Rev 1993, 12, 39–86. [Google Scholar]
  2. Drider, D.; Fimland, G.; Héchard, Y.; McMullen, L.M.; Prévost, H. The continuing story of class IIa bacteriocins. Microbiol. Mol. Biol. Rev 2006, 70, 564–582. [Google Scholar]
  3. Nissen-Meyer, J.; Rogne, P.; Oppegård, C.; Haugen, H.S.; Kristiansen, P.E. Structure-function relationships of the non-Lanthionine-containing peptide (class II) bacteriocins produced by Gram-positive bacteria. Curr. Pharm. Biotechnol 2009, 10, 19–37. [Google Scholar]
  4. Ennahar, S.; Sonomoto, K.; Ishizaki, A. Class IIa bacteriocins from lactic acid bacteria: Antibacterial activity and food preservation. J. Biosci. Bioeng 1999, 87, 705–716. [Google Scholar]
  5. Gálvez, A.; Abriouel, H.; López, R.L.; Ben Omar, N. Bacteriocin-based strategies for food biopreservation. Int. J. Food Microbiol 2007, 120, 51–70. [Google Scholar]
  6. García, P.; Rodríguez, L.; Rodríguez, A.; Martínez, B. Food biopreservation: Promising strategies using bacteriocins, bacteriophages and endolysins. Trends Food Sci. Technol 2010, 21, 373–382. [Google Scholar]
  7. Mills, S.; Stanton, C.; Hill, C.; Ross, R.P. New developments and applications of bacteriocins and peptides in foods. Annu. Rev. Food Sci. Technol 2011, 2, 299–329. [Google Scholar]
  8. Heng, N.C.K.; Burtenshaw, G.A.; Jack, R.W.; Tagg, J.R. Ubericin A, a Class IIa bacteriocin produced by Streptococcus uberis. Appl. Environ. Microbiol 2007, 73, 7763–7766. [Google Scholar]
  9. Nicolas, G.G.; LaPointe, G.; Lavoie, M.C. Production, purification, sequencing and activity spectra of mutacins D-123.1 and F-59.1. BMC Microbiol 2011, 11, 69. [Google Scholar]
  10. Yildirim, Z.; Johnson, M.G. Characterization and antimicrobial spectrum of bifidocin B, a bacteriocin produced by Bifidobacterium bifidum NCFB 1454. J. Food Prot 1998, 61, 47–51. [Google Scholar]
  11. Yildirim, Z.; Winters, D.K.; Johnson, M.G. Purification, amino acid sequence and mode of action of bifidocin B produced by Bifidobacterium bifidum NCFB 1454. J. Appl. Microbiol 1999, 86, 45–54. [Google Scholar]
  12. Cheikhyoussef, A.; Cheikhyoussef, N.; Chen, H.Q.; Zhao, J.X.; Tang, J.; Zhang, H. Bifidin I—A new bacteriocin produced by Bifidobacterium infantis BCRC 14602: Purification and partial amino acid sequence. Food Control 2010, 21, 746–753. [Google Scholar]
  13. Le Marrec, C.; Hyronimus, B.; Bressollier, P.; Verneuil, B.; Urdaci, M.C. Biochemical and genetic characterization of coagulin, a new antilisterial bacteriocin in the pediocin family of bacteriocins, produced by Bacillus coagulans I4. Appl. Environ. Microbiol 2000, 66, 5213–5220. [Google Scholar]
  14. Kalmokoff, M.L.; Banerjee, S.K.; Cyr, T.; Hefford, M.A.; Gleeson, T. Identification of a new plasmid-encoded sec-dependent bacteriocin produced by Listeria innocua 743. Appl. Environ. Microbiol 2001, 67, 4041–4047. [Google Scholar]
  15. Håvarstein, L.S.; Diep, D.B.; Nes, I.F. A family of bacteriocin ABC transporters carry out proteolytic processing of their substrates concomitant with export. Mol. Microbiol 1995, 16, 229–240. [Google Scholar]
  16. Zouhir, A.; Hammami, R.; Fliss, I.; Hamida, J.B. A new structure-based classification of Gram-positive bacteriocins. Protein J 2010, 29, 432–439. [Google Scholar]
  17. Rea, M.C.; Ross, R.P.; Cotter, P.D.; Hill, C. Classification of Bacteriocins from Gram-Positive Bacteria. In Prokaryotic Antimicrobial Peptides: From Genes to Applications; Drider, D., Rebuffat, S., Eds.; Springer Publishing Inc: New York, NY, USA, 2011; pp. 29–53. [Google Scholar]
  18. Diep, D.B.; Nes, I.F. Ribosomally synthesized antibacterial peptides in Gram positive bacteria. Curr. Drug Targets 2002, 3, 107–122. [Google Scholar]
  19. Belguesmia, Y.; Naghmouchi, K.; Chihib, N.-E.; Drider, D. Class IIa Bacteriocins: Current Knowledge and Perspectives. In Prokaryotic Antimicrobial Peptides: From Genes to Applications; Drider, D., Rebuffat, S., Eds.; Springer Publishing Inc.: New York, NY, USA, 2011; pp. 171–195. [Google Scholar]
  20. Fimland, G.; Johnsen, L.; Dalhus, B.; Nissen-Meyer, J. Pediocinlike antimicrobial peptides (class IIa bacteriocins) and their immunity proteins: Biosynthesis, structure and mode of action. J. Pept. Sci 2005, 11, 688–696. [Google Scholar]
  21. Birri, D.J.; Brede, D.A.; Forberg, T.; Holo, H.; Nes, I.F. Molecular and genetic characterization of a novel bacteriocin locus in Enterococcus avium isolates from infants. Appl. Environ. Microbiol 2010, 76, 483–492. [Google Scholar]
  22. Larsen, A.G.; Vogensen, F.K.; Josephsen, J. Antimicrobial activity of lactic acid bacteria isolated from sour doughs: Purification and characterization of bavaricin A, a bacteriocin produced by Lactobacillus bavaricus MI401. J. Appl. Microbiol 1993, 75, 113–122. [Google Scholar]
  23. Xiraphi, N.; Georgalaki, M.; van Driessche, G.; Devreese, B.; van Beeumen, J.; Tsakalidou, E.; Metaxopoulos, J.; Drosinos, E.H. Purification and characterization of curvaticin L442, a bacteriocin produced by Lactobacillus curvatus L442. Antonie Van Leeuwenhoek 2006, 89, 19–26. [Google Scholar]
  24. Saavedra, L.; Minahk, C.; de Ruiz Holgado, A.P.; Sesma, F. Enhancement of the enterocin CRL35 activity by a synthetic peptide derived from the NH2-terminal sequence. Antimicrob. Agents Chemother 2004, 48, 2778–2781. [Google Scholar]
  25. Papagianni, M.; Papamichae, E.M. Purification, amino acid sequence and characterization of the class IIa bacteriocin weissellin A, produced by Weissella paramesenteroides DX. Bioresour. Technol 2011, 102, 6730–6734. [Google Scholar]
  26. Svetoch, E.A.; Stern, N.J.; Eruslanov, B.V.; Kovalev, Y.N.; Volodina, L.I.; Perelygin, V.V.; Mitsevich, E.V.; Mitsevich, I.P.; Pokhilenko, V.D.; Borzenkov, V.N.; et al. Isolation of Bacillus circulans and Paenibacillus polymyxa strains inhibitory to Campylobacter jejuni and characterization of associated bacteriocins. J. Food Prot 2005, 68, 11–17. [Google Scholar]
  27. Kaiser, A.L.; Montville, T.J. Purification of the bacteriocin bavaricin MN and characterization of its mode of action against Listeria monocytogenes Scott A cells and lipid vesicles. J. Appl. Microbiol 1996, 62, 4529–4535. [Google Scholar]
  28. Svetoch, E.A.; Eruslanov, B.V.; Perelygin, V.V.; Mitsevich, E.V.; Mitsevich, I.P.; Borzenkov, V.N.; Levchuk, V.P.; Svetoch, O.E.; Kovalev, Y.N.; Stepanshin, Y.G.; et al. Diverse antimicrobial killing by Enterococcus faecium E 50–52 bacteriocin. J. Agric. Food Chem 2008, 56, 1942–1948. [Google Scholar]
  29. Kanatani, K.; Oshimura, M.; Sano, K. Isolation and characterization of acidocin A and cloning of the bacteriocin gene from Lactobacillus acidophilus. Appl. Environ. Microbiol 1995, 61, 1061–1067. [Google Scholar]
  30. Stern, N.J.; Svetoch, E.A.; Eruslanov, B.V.; Perelygin, V.V.; Mitsevich, E.V.; Mitsevich, I.P.; Pokhilenko, V.D.; Levchuk, V.P.; Svetoch, O.E.; Seal, B.S. Isolation of a Lactobacillus salivarius strain and purification of its bacteriocin, which is inhibitory to Campylobacter jejuni in the chicken gastrointestinal system. Antimicrob. Agents Chemother. 2006, 50, 3111–3116. [Google Scholar]
  31. Svetoch, E.A.; Eruslanov, B.V.; Levchuk, V.P.; Perelygin, V.V.; Mitsevich, E.V.; Mitsevich, I.P.; Stepanshin, J.; Dyatlov, I.; Seal, B.S.; Stern, N.J. Isolation of Lactobacillus salivarius 1077 (NRRL B-50053) and characterization of its bacteriocin, including the antimicrobial activity spectrum. Appl. Environ. Microbiol 2011, 77, 2749–2754. [Google Scholar]
  32. Feng, G.; Guron, G.K.; Churey, J.J.; Worobo, R.W. Characterization of mundticin L, a class IIa anti-Listeria bacteriocin from Enterococcus mundtii CUGF08. Appl. Environ. Microbiol 2009, 75, 5708–5713. [Google Scholar]
  33. Felix, J.V.; Papathanasopoulos, M.A.; Smith, A.A.; von Holy, A.; Hastings, J.W. Characterization of leucocin B-Ta11a, a bacteriocin from Leuconostoc carnosum Ta11a isolated from meat. Curr. Microbiol 1994, 29, 207–212. [Google Scholar]
  34. Flannagan, S.E.; Clewell, D.B.; Sedgley, C.M. A “retrocidal” plasmid in Enterococcus faecalis, passage and protection. Plasmid 2008, 59, 217–230. [Google Scholar]
  35. Arnold, K.; Bordoli, L.; Kopp, J.; Schwede, T. The SWISS-MODEL Workspace: A web-based environment for protein structure homology modelling. Bioinformatics 2006, 22, 195–201. [Google Scholar]
  36. Schwede, T.; Kopp, J.; Guex, N.; Peitsch, M.C. SWISS-MODEL: An automated protein homology-modeling server. Nucleic Acids Res 2003, 31, 3381–3385. [Google Scholar]
  37. Guex, N.; Peitsch, M.C. SWISS-MODEL and the Swiss-PdbViewer: An environment for comparative protein modelling. Electrophoresis 1997, 18, 2714–2723. [Google Scholar]
  38. Tichaczek, P.S.; Nissen-Meyer, J.; Nes, I.F.; Vogel, R.F.; Hammes, W.P. Characterization of the bacteriocins curvacin A from Lactobacillus curvatus LTH1174 and sakacin P from L. sake LTH673. Syst. Appl. Microbiol 1992, 15, 460–468. [Google Scholar]
  39. Cocolin, L.; Rantsiou, K. Sequencing and expression analysis of sakacin genes in Lactobacillus curvatus strains. Appl. Environ. Microbiol 2007, 76, 1403–1411. [Google Scholar]
  40. Tichaczek, P.S.; Vogel, R.F.; Hammes, W.P. Cloning and sequencing of sakP encoding sakacin P, the bacteriocin produced by Lactobacillus sake LTH 673. Microbiology 1994, 140, 361–367. [Google Scholar]
  41. Urso, R.; Rantsiou, K.; Cantoni, C.; Comi, G.; Cocolin, L. Sequencing and expression analysis of the sakacin P bacteriocin produced by a Lactobacillus sakei strain isolated from naturally fermented sausages. Appl. Environ. Microbiology 2006, 71, 480–485. [Google Scholar]
  42. Gonzalez, C.F.; Kunka, B.S. Plasmid-associated bacteriocin production and sucrose fermentation in Pediococcus acidilactici. Appl. Environ. Microbiol 1987, 53, 2534–2538. [Google Scholar]
  43. Bhunia, A.K.; Johnson, M.C.; Ray, B. Direct detection of an antimicrobial peptide of Pediococcus acidilactici in SDS-PAGE. J. Ind. Microbiol 1987, 2, 319–322. [Google Scholar]
  44. Motlagh, A.M.; Bhunia, A.K.; Szostek, F.; Hansen, T.R.; Johnson, M.C.; Ray, B. Nucleotide and amino acid sequence of pap-gene (pediocin AcH production) in Pediococcus acidilactici H. Lett. Appl. Microbiol 1992, 15, 45–48. [Google Scholar]
  45. Ray, S.K.; Johnson, M.C.; Ray, B. Bacteriocin plasmids of Pediococcus acidilactici. J. Ind. Microbiol 1989, 4, 163–171. [Google Scholar]
  46. Kim, W.J.; Ray, B.; Johnson, M.C. Plasmid transfers by conjugation and electroporation in Pediococcus acidilactici. J. Appl. Bacteriol 1992, 72, 201–207. [Google Scholar]
  47. Kwon, D.Y.; Koo, M.; Ryoo, C.R.; Kang, C.H.; Min, K.H.; Kim, W.J. Bacteriocin produced by Pediococcus sp. in kimchi and its characteristics. J. Microbiol. Biot 2002, 12, 96–105. [Google Scholar]
  48. Albano, H.; Todorov, S.D.; van Reenen, C.A.; Hogg, T.; Dicks, L.M.T.; Teixeira, P. Characterization of two bacteriocins produced by Pediococcus acidilactici isolated from “Alheira”, a fermented sausage traditionally produced in Portugal. Int. J. Food Microbiol 2007, 116, 239–247. [Google Scholar]
  49. Millette, M.; Dupont, C.; Shareck, F.; Ruiz, M.T.; Archambault, D.; Lacroix, M. Purification and identification of the pediocin produced by Pediococcus acidilactici MM33, a new human intestinal strain. J. Appl. Microbiol 2008, 104, 269–275. [Google Scholar]
  50. Bennik, M.H.J.; Verheul, A.; Abee, T.; Naaktgeboren-Stoffels, G.; Gorris, L.G.M.; Smid, E.J. Interactions of nisin and pediocin PA-1 with closely related lactic acid bacteria that manifest over 100-fold differences in bacteriocin sensitivity. Appl. Environ. Microbiol 1997, 63, 3628–3636. [Google Scholar]
  51. Casadei, G.; Grilli, E.; Piva, A. Pediocin A modulates intestinal microflora metabolism in swine in vitro intestinal fermentations. J. Anim. Sci 2009, 87, 2020–2028. [Google Scholar]
  52. Ennahar, S.; Aoude-Werner, D.; Sorokine, O.; van Dorsselaer, A.; Bringel, F.; Hubert, J.C.; Hasselmann, C. Production of pediocin AcH by Lactobacillus plantarum WHE92 isolated from cheese. Appl. Environ. Microbiol 1996, 62, 4381–4387. [Google Scholar]
  53. Bernbom, N.; Licht, T.R.; Saadbye, P.; Vogensen, F.K.; Norrung, B. Lactobacillus plantarum inhibits growth of Listeria monocytogenes in an in vitro continuous flow gut model, but promotes invasion of L. monocytogenes in the gut of gnotobiotic rats. Int. J. Food Microbiol 2006, 108, 10–14. [Google Scholar]
  54. Hoover, D.G.; Walsh, P.M.; Kolaetis, K.M.; Daly, M.M. A bacteriocin produced by Pediococcus species associated with a 5.5 megadalton plasmid. J. Food Prot 1988, 59, 29–31. [Google Scholar]
  55. Jager, K.; Harlander, S. Characterization of a bacteriocin from Pediococcus acidilactici PC and comparison of bacteriocin-producing strains using molecular typing procedures. Appl. Environ. Microbiol 1992, 37, 631–637. [Google Scholar]
  56. Ray, B.; Motlagh, A.M.; Johnson, M.C.; Bozoglu, F. Mapping of pSMB74, a plasmid encoding bacteriocin AcH production (Pap+) trait in Pediococcus acidilactici H. Lett. Appl. Microbiol 1992, 15, 35–37. [Google Scholar]
  57. Schved, F.; Lalazar, A.; Henis, Y.; Juven, B.J. Purification, partial characterization and plasmid linkage of pediocin SJ-1, a bacteriocin produced by Pediococcus acidilactici. J. Appl. Bacteriol 1993, 74, 67–77. [Google Scholar]
  58. Bhunia, A.K.; Bhowmik, T.K.; Johnson, M.G. Determination of bacteriocin-encoding plasmids of Pediococcus acidilactici strains by southern hybridization. Lett. Appl. Microbiol 1994, 18, 168–170. [Google Scholar]
  59. Rodríguez, J.M.; Cintas, L.M.; Casaus, P.; Martínez, M.I.; Suárez, A.; Hernández, P.E. Detection of pediocin PA-1 producing pediococci by rapid molecular producing by rapid molecular biology techniques. Food Microbiol 1997, 14, 363–371. [Google Scholar]
  60. Ray, S.K.; Kim, W.J.; Johnson, M.C.; Ray, B. Conjugal transfer of a plasmid encoding bacteriocin production and immunity in Pediococcus acidilactici H. J. Appl. Bacteriol 1989, 66, 393–399. [Google Scholar]
  61. Rodríguez, J.M.; Martínez, M.I.; Kok, J. Pediocin PA-1, a wide-spectrum bacteriocin from lactic acid bacteria. Crit. Rev. Food Sci. Nutr 2002, 42, 91–121. [Google Scholar]
  62. Miller, K.W.; Ray, P.; Steinmetz, T.; Hanekamp, T.; Ray, B. Gene organization and sequences of pediocin AcH/PA-1, production operons in Pediococcus and Lactococcus plasmids. Lett. Appl. Microbiol 2005, 40, 52–62. [Google Scholar]
  63. Meijer, W.J.J.; Wisman, G.B.A.; Terpstra, P.; Thorsted, P.B.; Thomas, C.M.; Holsappel, S.; Venema, G.; Bron, S. Rolling-circle plasmids from B. subtilis: Complete nucleotide sequences and analyses of genes of pTA1015, pTA1040, pTA1050 and pTA1060, and comparisons with related plasmids from Gram-positive bacteria. FEMS Microbiol. Rev 1998, 21, 337–368. [Google Scholar]
  64. Nicolas, G.G.; Lavoie, M.C.; Lapointe, G. Molecular genetics, genomics and biochemistry of mutacins. Genes Genomes Genomics 2007, 1, 193–208. [Google Scholar]
  65. Collado, M.C.; Hernández, M.; Sanz, Y. Production of bacteriocin-like compounds by human faecal Bifidobacterium strains. J. Food Prot 2005, 68, 1034–1040. [Google Scholar]
  66. Bernbom, N.; Jelle, B.; Brogren, C.H.; Vogensen, F.K.; Norrung, B.; Licht, T.R. Pediocin PA-1 and a pediocin producing Lactobacillus plantarum strain do not change the HMA rat microbiota. Int. J. Food Microbiol 2009, 130, 251–257. [Google Scholar]
  67. Fimland, G.; Sletten, K.; Nissen-Meyer, J. The complete aminoacid sequence of the pediocin-like antimicrobial peptide leucocin C. Biochem. Biophys. Res. Commun 2002, 295, 826–827. [Google Scholar]
  68. Papathanasopoulos, M.A.; Krier, F.; Revol-Junelles, A.-M.; Lefebvre, G.; le Caer, J.P.; von Holy, A.; Hastings, J.W. Multiple bacteriocin production by Leuconostoc mesenteroides TA33a and other Leuconostoc/Weissella strains. Curr. Microbiol 1997, 35, 331–335. [Google Scholar]
  69. Papathanasopoulos, M.A.; Dykes, G.A.; Revol-Junelles, A.-M.; Delfour, A.; von Holy, A.; Hastings, J.W. Sequence and structural relationships of leucocins A-, B- and C-TA33a from Leuconostoc mesenteroides TA33a. Microbiology 1998, 144, 1343–1348. [Google Scholar]
  70. Sánchez, J.; Die, D.B.; Herranz, C.; Nes, I.F.; Cintas, L.M.; Hernandez, P.E. Amino acid and nucleotide sequence, adjacent genes, and heterologous expression of hiracin JM79, a sec-dependent bacteriocin produced by Enterococcus hirae DCH5, isolated from Mallard ducks (Anas platyrhynchos). FEMS Microbiol. Lett 2007, 270, 227–236. [Google Scholar]
  71. Doi, K.; Eguchi, T.; Choi, S.-H.; Iwatake, A.; Ohmomo, S.; Ogata, S. Isolation of enterocin SE-K4-encoding plasmid and a high enterocin SE-K4 producing strain of Enterococcus faecalis K-4. J. Biosci. Bioeng 2002, 93, 434–436. [Google Scholar]
  72. Tomita, H.; Fujimoto, S.; Tanimoto, K.; Ike, Y. Cloning and genetic organization of the bacteriocin 31 determinant encoded on the Enterococcus faecalis pheromone-responsive conjugative plasmid pYI17. J. Bacteriol 1996, 178, 3585–3593. [Google Scholar]
  73. Van Reenen, C.A.; Dicks, L.M.T.; Chikindas, M.L. Isolation, purification and partial characterization of plantaricin 423, a bacteriocin produced by Lactobacillus plantarum. J. Appl. Microbiol 1998, 84, 1131–1137. [Google Scholar]
  74. Gallager, N.L.F.; Sailer, V.; Niemczura, W.P.; Nakashima, T.T.; Stiles, M.E.; Vederas, J.C. Three-dimensional structure of leucocin A in trifluoroethanol and dodecylphosphocholine micelles: Spatial location of residues critical for biological activity in type IIa bacteriocins from lactic acid bacteria. Biochemistry 1997, 36, 15062–15072. [Google Scholar]
  75. Sit, C.S.; Lohans, C.T.; van Belkum, M.J.; Campbell, C.D.; Miskolzie, M.; Vederas, J.C. Substitution of a conserved disulfide in the type IIa bacteriocin, leucocin A, with l-leucine and l-serine residues: Effects on activity and three-dimensional structure. ChemBioChem 2012, 13, 35–38. [Google Scholar]
  76. Ferchichi, M.; Frère, J.; Mabrouk, K.; Manai, M. Lactococcin MMFII, a novel class IIa bacteriocin produced by Lactococcus lactis MMFII, isolated from a Tunisian dairy product. FEMS Microbiol. Lett 2001, 205, 49–55. [Google Scholar]
  77. Quadri, L.E.; Sailer, M.; Roy, K.L.; Vederas, J.C.; Stiles, M.E. Chemical and genetic characterization of bacteriocins produced by Carnobacterium piscicola LV17B. J. Biol. Chem 1994, 269, 12204–12211. [Google Scholar]
  78. Verluyten, J.; Messens, W.; de Vuyst, L. Sodium chloride reduces production of Curvacin A, a bacteriocin produced by Lactobacillus curvatus strain LTH 1174, originating from fermented sausage. Appl. Environ. Microbiol 2004, 70, 2271–2278. [Google Scholar]
  79. Cintas, L.M.; Casaus, P.; Havarstein, L.S.; Hernandez, P.E.; Nes, I.F. Biochemical and genetic characterization of enterocin P, a novel sec-dependent bacteriocin from Enterococcus faecium P13 with a broad antimicrobial spectrum. Appl. Environ. Microbiol 1997, 63, 4321–4330. [Google Scholar]
  80. Park, S.H.; Itoh, K.; Fujisawa, T. Characteristics and identification of enterocins produced by Enterococcus faecium JCM 5804T. J. Appl. Microbiol 2003, 95, 294–300. [Google Scholar]
  81. Kang, J.H.; Lee, M.S. Characterization of a bacteriocin produced by Enterococcus faecium GM-1 isolated from an infant. J. Appl. Microbiol 2005, 98, 1169–1176. [Google Scholar]
  82. Samuel, A.; Calo, P.; Franco, C.M.; Prado, M.; Cepeda, A.; Barros-Velazquez, J. Single nucleotide polymorphism analysis of the enterocin P structural gene of Enterococcus faecium strains isolated from nonfermented animal foods. Mol. Nutr. Food Res 2006, 20, 1229–1238. [Google Scholar]
  83. Hosseini, S.V.; Arlindo, S.; Böhme, K.; Fernández-No, C.; Calo-Mata, P.; Barros-Velázquez, J. Molecular and probiotic characterization of bacteriocin-producing Enterococcus faecium strains isolated from nonfermented animal foods. J. Appl. Microbiol 2009, 107, 1392–1403. [Google Scholar]
  84. Javed, I.; Ahmed, S.; Manam, S.; Riaz, M.; Ahmad, B.; Ali, M.I.; Hameed, A.; Chaudry, G.J. Production, characterization, and antimicrobial activity of a bacteriocin from newly isolated Enterococcus faecium IJ-31. J. Food Prot 2010, 73, 44–52. [Google Scholar]
  85. Waters, C.M.; Bassler, B.L. Quorum sensing: Cell-to-cell communication in bacteria. Annu. Rev. Cell Dev. Biol 2005, 21, 319–346. [Google Scholar]
  86. Kleerebezem, M.; Quadri, L.E.N.; Kuipers, O.P.; de Vos, W.M. Quorum sensing by peptide pheromones and two-component signal-transduction systems in Gram-positive bacteria. Mol. Microbiol 1997, 24, 895–904. [Google Scholar]
  87. Atkinson, S.; Williams, P. Quorum sensing and social networking in the microbial world. J. R. Soc. Interface 2009, 6, 959–978. [Google Scholar]
  88. Nes, I.F.; Eijsink, V.G.H. Regulation of Group II Peptide Bacteriocin Synthesis by Quorum-Sensing Mechanisms. In Cell-Cell Signalling in Bacteria; Dunny, G.M., Winans, S.C., Eds.; American Society for Microbiology: Washington, DC, USA, 1999; pp. 175–192. [Google Scholar]
  89. Ennahar, S.; Sashihara, T.; Sonomoto, K.; Ishizaki, A. Class IIa bacteriocins: Biosynthesis, structure and activity. FEMS Microbiol. Rev 2000, 24, 85–106. [Google Scholar]
  90. Cho, H.S.; Pelton, J.G.; Yan, D.; Kustu, S.; Wemmer, D.E. Phosphoaspartates in bacterial signal transduction. Curr. Opin. Struct. Biol 2001, 11, 679–684. [Google Scholar]
  91. Aymerich, T.; Holo, H.; Havarstein, L.S.; Hugas, M.; Garriga, M.; Nes, I.F. Biochemical and genetic characterization of enterocin A from Enterococcus faecium, a new antilisterial bacteriocin in the pediocin family of bacteriocins. Appl. Environ. Microbiol 1996, 62, 1676–1682. [Google Scholar]
  92. Métivier, A.; Pilet, M.-F.; Dousset, X.; Sorokine, O.; Anglade, P.; Zagorec, M.; Piard, J.-C.; Marion, D.; Cenatiempo, Y.; Fremaux, C. Divercin V41, a new bacteriocin with two disulphide bonds produced by Carnobacterium divergens V41: Primary structure and genomic organization. Microbiology 1998, 144, 2837–2844. [Google Scholar]
  93. Hühne, K.; Axelsson, L.; Holck, A.; Kröckel, L. Analysis of the sakacin P gene cluster from Lactobacillus sake Lb674 and its expression in sakacin-negative Lb. sake strains. Microbiology 1996, 142, 1437–1448. [Google Scholar]
  94. Quadri, L.E.N.; Kleerebezem, M.; Kuipers, O.P.; de Vos, W.M.; Roy, K.L.; Vederas, J.C.; Stiles, M.E. Characterization of a locus from Carnobacterium piscicola LV17B involved in bacteriocin production and immunity: Evidence for global inducer-mediated transcriptional regulation. J. Bacteriol 1997, 179, 6163–6171. [Google Scholar]
  95. Herranz, C.; Driessen, A.J.M. Sec-mediated secretion of bacteriocin enterocin P by Lactococcus lactis. Appl. Environ. Microbiol 2005, 71, 1959–1963. [Google Scholar]
  96. Gierasch, L.M. Signal sequences. Biochemistry 1989, 28, 923–930. [Google Scholar]
  97. Pugsley, A.P. The complete general secretory pathway in Gram-negative bacteria. Microbiol. Rev 1993, 57, 50–108. [Google Scholar]
  98. Izard, J.W.; Kendall, D.A. Signal peptides: Exquisitely designed transport promoters. Microbiol. Biotechnol 1994, 13, 765–773. [Google Scholar]
  99. Van Belkum, M.J.; Stiles, M.E. Molecular characterization of genes involved in the production of the bacteriocin leucocin A from Leuconostoc gelidum. Appl. Environ. Microbiol 1995, 61, 3573–3579. [Google Scholar]
  100. Nes, I.F.; Diep, D.B.; Håvarstein, L.S.; Brurberg, M.B.; Eijsink, V.; Holo, H. Biosynthesis of bacteriocins in lactic acid bacteria. Antonie Van Leeuwenhoek 1996, 70, 113–128. [Google Scholar]
  101. Vaughan, A.; Eijsink, V.G.; van Sinderen, D. Functional characterization of a composite bacteriocin locus from malt isolate Lactobacillus sakei 5. Appl. Environ. Microbiol 2003, 69, 7194–7203. [Google Scholar]
  102. Revol-Junelles, A.M.; Mathis, R.; Krier, F.; Fleury, Y.; Delfour, A.; Lefebvre, G. Leuconostoc mesenteroides subsp. mesenteroides FR52 synthesizes two distinct bacteriocins. Lett. Appl. Microbiol 1996, 23, 120–124. [Google Scholar]
  103. Fremaux, C.; Héchard, Y.; Cenatiempo, Y. Mesentericin Y105 gene clusters in Leuconostoc rnesenteroides Y 105. Microbiology 1995, 14, 1637–1645. [Google Scholar]
  104. Aucher, W.; Simonet, V.; Fremaux, C.; Dalet, K.; Simon, L.; Cenatiempo, Y.; Frère, J.; Berjeaud, J.M. Differences in mesentericin secretion systems from two Leuconostoc strains. FEMS Microbiol. Lett 2004, 232, 15–22. [Google Scholar]
  105. Simon, L.; Fremaux, C.; Cenatiempo, Y.; Berjeaud, J.M. Sakacin G, a new type of antilisterial bacteriocin. Appl. Environ. Microbiol 2002, 68, 6416–6420. [Google Scholar]
  106. Dortu, C.; Huch, M.; Holzapfel, W.H.; Franz, C.M.A.P.; Thonart, P. Anti-listerial activity of bacteriocin-producing Lactobacillus curvatus CWBI-B28 and Lactobacillus sakei CWBI-B1365 on raw beef and poultry meat. Lett. Appl. Microbiol 2008, 47, 581–586. [Google Scholar]
  107. Todorov, S.D.; Rachman, C.; Fourrier, A.; Dicks, L.M.T.; van Reenen, C.A.; Prévost, H.; Dousset, X. Characterization of a bacteriocin produced by Lactobacillus sakei R1333 isolated from smoked salmon. Anaerobe 2011, 17, 23–31. [Google Scholar]
  108. Wang, Y.; Henz, M.E.; Gallagher, N.L.; Chai, S.; Gibbs, A.C.; Yan, L.Z.; Stiles, M.E.; Wishart, D.S.; Vederas, J.C. Solution structure of carnobacteriocin B2 and implications for structure-activity relationships among type IIa bacteriocins from lactic acid bacteria. Biochemistry 1999, 38, 15438–15447. [Google Scholar]
  109. Uteng, M.; Hauge, H.H.; Markwick, P.R.; Fimland, G.; Mantzilas, D.; Nissen-Meyer, J.; Muhle-Goll, C. Three-dimensional structure in lipid micelles of the pediocin-like antimicrobial peptide sakacin P and a sakacin P variant that is structurally stabilized by an inserted C-terminal disulfide bridge. Biochemistry 2003, 42, 11417–11426. [Google Scholar]
  110. Haugen, H.S.; Fimland, G.; Nissen-Meyer, J.; Kristiansen, P.E. Three-dimensional structure in lipid micelles of the pediocin-like antimicrobial peptide curvacin A. Biochemistry 2005, 44, 16149–16157. [Google Scholar]
  111. Kaur, K.; Andrew, L.C.; Wishart, D.S.; Vederas, J.C. Dynamic relationships among type IIa bacteriocins: Temperature effects on antimicrobial activity and on structure of the C-terminal amphipathic α-helix as a receptor binding region. Biochemistry 2004, 43, 9009–9020. [Google Scholar]
  112. Fimland, G.; Blingsmo, O.R.; Sletten, K.; Jung, G.; Nes, I.F.; Nissen-Meyer, J. New biologically active hybrid bacteriocins constructed by combining regions from various pediocin-like bacteriocins: The C-terminal region is important for determining specificity. Appl. Environ. Microbiol 1996, 62, 3313–3318. [Google Scholar]
  113. Fimland, G.; Johnsen, L.; Axelsson, L.; Brurberg, M.B.; Nes, I.F.; Eijsink, V.G.H.; Nissen-Meyer, J. A C-terminal disulfide bridge in pediocin-like bacteriocins renders bacteriocin activity less temperature dependent and is a major determinant of the antimicrobial spectrum. J. Bacteriol 2000, 182, 2643–2648. [Google Scholar]
  114. Derksen, D.J.; Boudreau, M.A.; Vederas, J.C. Hydrophobic interactions as substitutes for a conserved disulfide linkage in the type IIa bacteriocins, leucocin A and pediocin PA-1. ChemBioChem 2008, 9, 1898–1901. [Google Scholar]
  115. Fleury, Y.; Dayem, M.A.; Montagne, J.J.; Chaboisseau, E.; le Caer, J.P.; Nicolas, P.; Delfour, A. Covalent structure, synthesis, and structure-function studies of mesentericin Y 105(37), a defensive peptide from gram-positive bacteria Leuconostoc mesenteroides. J. Biol. Chem 1996, 271, 14421–14429. [Google Scholar]
  116. Tominaga, T.; Hatakeyama, Y. Determination of essential and variable residues in pediocin PA-1 by NNK scanning. Appl. Environ. Microbiol 2006, 72, 1141–1147. [Google Scholar]
  117. Derksen, D.J.; Stymiest, J.L.; Vederas, J.C. Antimicrobial leucocin analogues with a disulfide bridge replaced by a carbocycle or by noncovalent interactions of allyl glycine residues. J. Am. Chem. Soc 2006, 128, 14252–14253. [Google Scholar]
  118. Hutchinson, E.G.; Thornton, J.M. A revised set of potentials for beta-turn formation in proteins. Protein Sci 1994, 3, 2207–2216. [Google Scholar]
  119. Yan, L.Z.; Gibbs, A.C.; Stiles, M.E.; Wishart, D.S.; Vederas, J.C. Analogues of bacteriocins: antimicrobial specificity and interactions of leucocin A with its enantiomer, carnobacteriocin B2, and truncated derivatives. J. Med. Chem 2000, 43, 4579–4581. [Google Scholar]
  120. Quadri, L.E.N.; Yan, L.Z.; Stiles, M.E.; Vederas, J.C. Overproduction of the antimicrobial peptide, its engineered variants, and its precursor in Escherichia coli. J. Biol. Chem 1997, 272, 3384–3388. [Google Scholar]
  121. Fimland, G.; Jack, R.; Jung, G.; Nes, I.F.; Nissen-Meyer, J. The bactericidal activity of pediocin PA-1 is specifically inhibited by a 15-mer fragment that spans the bacteriocin from the center toward the C terminus. Appl. Environ. Microbiol 1998, 64, 5057–5060. [Google Scholar]
  122. Eijsink, V.G.; Skeie, M.; Middelhoven, P.H.; Brurberg, M.B.; Nes, I.F. Comparative studies of class IIa bacteriocins of lactic acid bacteria. Appl. Environ. Microbiol 1998, 64, 3275–3281. [Google Scholar]
  123. Richard, C.; Cañon, R.; Naghmouchi, K.; Bertrand, D.; Prévosta, H.; Drider, D. Evidence on correlation between number ofdisulfide bridge and toxicity of class IIa bacteriocins. Food Microbiol 2006, 23, 175–183. [Google Scholar]
  124. Robichon, D.; Gouin, E.; Débarbouillé, M.; Cossart, P.; Cenatiempo, Y.; Héchard, Y. The rpoN54) gene from Listeria monocytogenes is involved in resistance to mesentericin Y105, an antibacterial peptide from Leuconostoc mesenteroides. J. Bacteriol 1997, 179, 7591–7594. [Google Scholar]
  125. Ramnath, M.; Beukes, M.; Tamura, K.; Hastings, J.W. Absence of a putative mannose-specific phosphotransferase system enzyme IIAB component in a leucocin A-resistant strain of Listeria monocytogenes, as shown by two-dimensional sodium dodecyl sulfatepolyacrylamide gel electrophoresis. Appl. Environ. Microbiol 2000, 66, 3098–3101. [Google Scholar]
  126. Dalet, K.; Briand, C.; Cenatiempo, Y.; Héchard, Y. The rpoN gene of Enterococcus faecalis directs sensitivity to subclass IIa bacteriocins. Curr. Microbiol 2000, 41, 441–443. [Google Scholar]
  127. Héchard, Y.; Pelletier, C.; Cenatiempo, Y.; Frère, J. Analysis of σ54-dependent genes in Enterococcus faecalis: A mannose PTS permease (EIIMan) is involved in sensitivity to a bacteriocin, mesentericin Y105. Microbiology 2001, 147, 1575–1580. [Google Scholar]
  128. Ramnath, M.; Arous, S.; Gravesen, A.; Hastings, J.W.; Héchard, Y. Expression of mptC of Listeria monocytogenes induces sensitivity to class IIa bacteriocins in Lactococcus lactis. Microbiology 2004, 150, 2663–2668. [Google Scholar]
  129. Diep, D.B.; Skaugen, M.; Salehian, Z.; Holo, H.; Nes, I.F. Common mechanisms of target cell recognition and immunity for class II bacteriocins. Proc. Natl. Acad. Sci. USA 2007, 104, 2384–2389. [Google Scholar]
  130. Kjos, M.; Salehian, Z.; Nes, I.F.; Diep, D.B. An extracellular loop of the mannose phosphotransferase system component IIC is responsible for specific targeting by class IIa bacteriocins. J. Bacteriol 2010, 192, 5906–5913. [Google Scholar]
  131. Kjos, M.; Borrero, J.; Opsata, M.; Birri, D.J.; Holo, H.; Cintas, L.M; Snipen, L.; Hernández, P.E; Nes, I.F.; Diep, D.B. Target recognition, resistance, immunity and genome mining of class II bacteriocins from Gram-positive bacteria. Microbiology 2011, 157, 3256–3267. [Google Scholar]
  132. Postma, P.W.; Lengeler, J.W.; Jacobson, G.R. Phosphoenolpyruvate: Carbohydrate phosphotransferase systems of bacteria. Microbiol. Rev 1993, 57, 543–594. [Google Scholar]
  133. Mao, Q.; Schunk, T.; Flukiger, K.; Erni, B. Functional reconstitution of the purified mannose phosphotransferase system of Escherichia coli into phospholipid vesicles. J. Biol. Chem 1995, 270, 5258–5265. [Google Scholar]
  134. Hassan, M.; Kjos, M.; Nes, I.F.; Diep, D.B.; Lotfipour, F. Natural antimicrobial peptides from bacteria: Characteristics and potential applications to fight against antibiotic resistance. J. Appl. Microbiol 2012, 113, 723–736. [Google Scholar]
  135. Kjos, M.; Nes, I.F.; Diep, D.B. Mechanisms of resistance to bacteriocins targeting the mannose phosphotransferase system. Appl. Environ. Microbiol. 2011, 77, 3335–3342. [Google Scholar]
  136. Kjos, M.; Nes, I.F.; Diep, D.B. Class II one-peptide bacteriocins target a phylogenetically defined subgroup of mannose phosphotransferase systems on sensitive cells. Microbiology 2009, 155, 2949–2961. [Google Scholar]
  137. Diep, D.B.; Godager, L.; Brede, D.; Nes, I.F. Data mining and characterization of a novel pediocin-like bacteriocin system from the genome of Pediococcus pentosaceus ATCC 25745. Microbiology 2006, 152, 1649–1659. [Google Scholar]
  138. Katla, T.; Naterstad, K.; Vancanneyt, M.; Swings, J.; Axelsson, L. Differences in susceptibility of Listeria monocytogenes strains to sakacin P, sakacin A, pediocin PA-1, and nisin. Appl. Environ. Microbiol 2003, 69, 4431–4437. [Google Scholar]
  139. Johnsen, L.; Fimland, G.; Nissen-Meyer, J. The C-terminal domain of pediocin-like antimicrobial peptides (class IIa bacteriocins) is involved in specific recognition of the C-terminal part of cognate immunity proteins and in determining the antimicrobial spectrum. J. Biol. Chem 2005, 280, 9243–9250. [Google Scholar]
  140. Haugen, H.S.; Fimland, G.; Nissen-Meyer, J. Mutational analysis of residues in the helical region of the class IIa bacteriocin pediocin PA-1. Appl. Environ. Microbiol 2011, 77, 1966–1972. [Google Scholar]
  141. Erni, B. The mannose transporter complex: An open door for the macromolecular invasion of bacteria. J. Bacteriol 2006, 188, 7036–7038. [Google Scholar]
  142. Gravesen, A.; Ramnath, M.; Rechinger, K.B.; Andersen, N.; Jansch, L.; Héchard, Y.; Hastings, J.W.; Knøchel, S. High-level resistance to class IIa bacteriocins is associated with one general mechanism in Listeria monocytogenes. Microbiology 2002, 148, 2361–2369. [Google Scholar]
  143. Tessema, G.T.; Moretro, T.; Kohler, A.; Axelsson, L.; Naterstad, K. Complex phenotypic and genotypic responses of Listeria monocytogenes strains exposed to the class IIa bacteriocin sakacin P. Appl. Environ. Microbiol 2009, 75, 6973–6980. [Google Scholar]
  144. Dalet, K.; Cenatiempo, Y.; Cossart, P.; Hechard, Y. A sigma (σ54)-dependent PTS permease of the mannose family is responsible for sensitivity of Listeria monocytogenes to mesentericin Y105. Microbiology 2001, 147, 3263–3269. [Google Scholar]
  145. Duffes, F.; Jenoe, P.; Boyaval, P. Use of two-dimensional electrophoresis to study differential protein expression in divercin V41-resistant and wild-type strains of Listeria monocytogenes. Appl. Environ. Microbiol 2000, 66, 4318–4324. [Google Scholar]
  146. Xue, J.; Hunter, I.; Steinmetz, T.; Peters, A.; Ray, B.; Miller, K.W. Novel activator of mannose-specific phosphotransferase system permease expression in Listeria innocua, identified by screening for pediocin AcH resistance. Appl. Environ. Microbiol 2005, 71, 1283–1290. [Google Scholar]
  147. Arous, S.; Dalet, K.; Hechard, Y. Involvement of the mpo operon in resistance to class IIa bacteriocins in Listeria monocytogenes. FEMS Microbiol. Lett 2004, 238, 37–41. [Google Scholar]
  148. Kaur, G.; Malik, R.K.; Mishra, S.K.; Singh, T.P.; Bhardwaj, A.; Singroha, G.; Vij, S.; Kumar, N. Nisin and class IIa bacteriocin resistance among Listeria and other foodborne pathogens and spoilage bacteria. Microb. Drug Resist 2011, 17, 197–205. [Google Scholar]
  149. Simon, L.; Fremaux, C.; Cenatiempo, Y.; Berjeaud, J.-M. Luminescent method for the detection of antibacterial activities. Appl. Microbiol. Biotechnol 2001, 57, 757–763. [Google Scholar]
  150. Eijsink, V.G.H.; Axelsson, L.; Diep, D.B.; Håvarstein, L.S.; Holo, H.; Nes, I.F. Production of class II bacteriocins by lactic acid bacteria: An example of biological warfare and communication. Antonie Van Leeuwenhoek 2002, 81, 639–654. [Google Scholar]
  151. Rouse, S.; Sun, F.; Vaughan, A.; van Sinderen, D. High-throughput isolation of bacteriocin-producing lactic acid bacteria, with potential application in the brewing industry. J. Inst. Brew 2007, 113, 256–262. [Google Scholar]
  152. Więckowicz, M.; Schmidt, M.; Sip, A.; Grajek, W. Development of a PCR-based assay for rapid detection of class IIa bacteriocin genes. Lett. Appl. Microbiol 2011, 52, 281–289. [Google Scholar]
  153. Losteinkit, C.; Uchiyama, K.; Ochi, S.; Takaoka, T.; Nagahisa, K.; Shioya, S. Characterization of bacteriocin N15 produced by Enterococcus faecium N15 and cloning of the related genes. J. Biosci. Bioeng 2001, 91, 390–395. [Google Scholar]
  154. Choho, G.; Abriouel, H.; Omar, N.; López, R.; Ortega, E.; Martínez-Cañamero, M.; Laglaoui, A.; Barrijal, S.; Gálvez, A. Characterization of a bacteriocin-producing strain of Enterococcus faecalis from cow’s milk used in the production of Moroccan traditional dairy foods. World J. Microbiol. Biotechnol 2008, 24, 997–1001. [Google Scholar]
  155. Belgacem, Z.B.; Abriouel, H.; Omar, N.B.; Lucas, R.; Martínez-Canamero, M.; Gálvez, A.; Manai, M. Antimicrobial activity, safety aspects, and some technological properties of bacteriocinogenic Enterococcus faecium from artisanal Tunisian fermented meat. Food Control 2010, 21, 462–470. [Google Scholar]
  156. Yi, H.; Zhang, L.; Tuo, Y.; Han, X.; Du, M. A novel method for rapid detection of class IIa bacteriocin-producing lactic acid bacteria. Food Control 2010, 21, 426–430. [Google Scholar]
  157. Knoll, C.; Divol, B.; Toit, M. Genetic screening of lactic acid bacteria of oenological origin for bacteriocin-encoding genes. Food Microbiol 2008, 25, 983–991. [Google Scholar]
  158. Liu, M.J.; van Enckevort, F.H.J.; Siezen, R.J. Genome update: Lactic acid bacteria genome sequencing is booming. Microbiology 2005, 151, 3811–3814. [Google Scholar]
  159. Pfeiler, E.A.; Klaenhammer, T.R. The genomics of lactic acid bacteria. Trends Microbiol 2007, 15, 546–553. [Google Scholar]
  160. Nes, I.F.; Johnsborg, O. Exploration of antimicrobial potential in LAB by genomics. Curr. Opin. Biotechnol 2004, 15, 100–104. [Google Scholar]
  161. Dirix, G.; Monsieurs, P.; Dombrecht, B.; Daniels, R.; Marchalb, K.; Vanderleydena, J.; Michielsa, J. Peptide signal molecules and bacteriocins in Gram-negative bacteria: A genome-wide in silico screening for peptides containing a double-glycine leader sequence and their cognate transporters. Peptides 2004, 25, 1425–1440. [Google Scholar]
  162. Dirix, G.; Monsieurs, P.; Marchal, K.; Vanderleyden, J.; Michiels, J. Screening genomes of Gram-positive bacteria for double-glycine-motif containing peptides. Microbiology 2004, 150, 1121–1126. [Google Scholar]
  163. Wang, Z.; Wang, G. APD: The antimicrobial peptide database. Nucleic Acids Res 2004, 32, D590–D592. [Google Scholar]
  164. Fjell, C.D.; Jenssen, H.; Hilpert, K.; Cheung, W.A.; Pante, N.; Hancock, R.E.; Cherkasov, A. Identification of novel antibacterial peptides by chemoinformatics and machine learning. J. Med. Chem 2009, 52, 2006–2015. [Google Scholar]
  165. Fjell, C.D.; Hancock, R.E.; Cherkasov, A. AMPer: A database and an automated discovery tool for antimicrobial peptides. Bioinformatics 2007, 23, 1148–1155. [Google Scholar]
  166. Bactibase: Database dedicated to bacteriocins, Available online: accessed on 20 November 2012.
  167. Bagel2: The bacteriocin mining tool, Available online: accessed on 20 November 2012.
  168. Hammami, R.; Zouhir, A.; Lay, C.L.; Hamida, J.B.; Fliss, I. BACTIBASE second release: A database and tool platform for bacteriocin characterization. BMC Microbiol 2010, 10, 22. [Google Scholar]
  169. De Jong, A.; van Heel, A.J.; Kok, J.; Kuipers, O.P. BAGEL2: Mining for bacteriocins in genomic data. Nucleic Acids Res 2010, 38, W647–W651. [Google Scholar]
  170. Wang, P.; Hu, L.; Liu, G.Y.; Jiang, N.; Chen, X.Y.; Xu, J.Y.; Zheng, W.; Li, L.; Tan, M.; Chen, Z.; et al. Prediction of antimicrobial peptides based on sequence alignment and feature selection methods. PLoS One 2011, 6, e18476. [Google Scholar]
  171. Fernandes, F.C.; Rigden, D.J.; Franco, O.L. Prediction of antimicrobial peptides based on the adaptive neuro-Fuzzy inference system application. Pept. Sci 2012, 98, 280–287. [Google Scholar]
  172. Hammami, R.; Fliss, I. Current trends in antimicrobial agent research: Chemo- and bioinformatics approaches. Drug Discov. Today 2010, 15, 540–546. [Google Scholar]
  173. Hilpert, K.; Elliott, M.R.; Volkmer-Engert, R.; Henklein, P.; Donini, O.; Zhou, Q.; Winkler, D.F.; Hancock, R.E. Sequence requirements and an optimization strategy for short antimicrobial peptides. Chem. Biol 2006, 13, 1101–1107. [Google Scholar]
  174. Jenssen, H.; Fjell, C.D.; Cherkasov, A.; Hancock, R.E. QSAR modeling and computer-aided design of antimicrobial peptides. J. Pept. Sci 2008, 14, 110–114. [Google Scholar]
  175. Frecer, V.; Ho, B.; Ding, J.L. De novo design of potent antimicrobial peptides. Antimicrob. Agents Chemother 2004, 48, 3349–3357. [Google Scholar]
  176. Cherkasov, A.; Jankovic, B. Application of ‘inductive’ QSAR descriptors for quantification of antibacterial activity of cationic polypeptides. Molecules 2004, 9, 1034–1052. [Google Scholar]
  177. Wang, Y.Q.; Ding, Y.; Wen, H.X.; Lin, Y.; Hu, Y.; Zhang, Y.; Xia, Q.Y.; Li, Z.H. QSAR modeling and design of cationic antimicrobial peptides based on structural properties of amino acids. Comb. Chem. High Throughput Screening 2012, 15, 347–353. [Google Scholar]
  178. Melo, M.N.; Ferre, R.; Feliu, L.; Bardají, E.; Planas, M.; Castanho, M.A.R.B. Prediction of antibacterial activity from physicochemical properties of antimicrobial peptides. PLoS One 2011, 6, e28549. [Google Scholar]
  179. Huang, H.W. Molecular mechanism of antimicrobial peptides: The origin of cooperativity. Biochim. Biophys. Acta 2006, 1758, 1292–1302. [Google Scholar]
  180. Leontiadou, H.; Mark, A.E.; Marrink, S.J. Antimicrobial peptides in action. J. Am. Chem. Soc 2006, 128, 12156–12161. [Google Scholar]
  181. Melo, M.N.; Castanho, M.A. Omiganan interaction with bacterial membranes and cell wall models. Assigning a biological role to saturation. Biochim. Biophys. Acta 2007, 1768, 1277–1290. [Google Scholar]
  182. Pistolesi, S.; Pogni, R.; Feix, J.B. Membrane insertion and bilayer perturbation by antimicrobial peptide CM15. Biophys. J 2007, 93, 1651–1660. [Google Scholar]
  183. Melo, M.N.; Ferre, R.; Castanho, M.A. Antimicrobial peptides: Linking partition, activity and high membrane-bound concentrations. Nat. Rev. Microbiol 2009, 7, 245–250. [Google Scholar]
  184. Alves, C.S.; Melo, M.N.; Franquelim, H.G.; Ferre, R.; Planas, M.; Feliu, L.; Bardají, E.; Kowalczyk, W.; Andreu, D.; Santos, N.C.; et al. Escherichia coli cell surface perturbation and disruption induced by antimicrobial peptides BP100 and pepR. J. Biol. Chem 2010, 285, 27536–27544. [Google Scholar]
  185. Acuña, L.; Morero, R.D.; Bellomio, A. Development of wide-spectrum hybrid bacteriocins for food biopreservation. Food Bioprocess Technol 2011, 4, 1029–1049. [Google Scholar]
  186. Kalmokoff, M.L.; Daley, E.; Austin, J.W.; Farber, J.M. Bacteriocin-like inhibitory activities among various species of Listeria. Int. J. Food Microbiol 1999, 50, 191–201. [Google Scholar]
  187. Bennik, M.H.J.; Vanloo, B.; Brasseur, R.; Gorris, L.G.M.; Smid, E.J. A novel bacteriocin with a YGNGV motif from vegetable-associated Enterococcus mundtii: full characterization and interaction with target organisms. Biochim. Biophys. Acta 1998, 1373, 47–58. [Google Scholar]
  188. Kawamoto, S.; Shima, J.; Sato, R.; Eguchi, T.; Ohmomo, S.; Shibato, J.; Horikoshi, N.; Takeshita, K.; Sameshima, T. Biochemical and genetic characterization of mundticin KS, an antilisterial peptide produced by Enterococcus mundtii NFRI 7393. J. Appl. Microbiol 2002, 68, 3830–3840. [Google Scholar]
  189. Zendo, T.; Eungruttanagorn, N.; Fujioka, S.; Tashiro, Y.; Nomura, K.; Sera, Y.; Kobayashi, G.; Nakayama, J.; Ishizaki, A.; Sonomoto, K. Identification and production of a bacteriocin from Enterococcus mundtii QU2 isolated from soybean. J. Appl. Microbiol 2005, 99, 1181–1190. [Google Scholar]
  190. Yamazaki, K.; Suzuki, M.; Kawai, Y.; Inoue, N.; Montville, T.J. Purification and characterization of a novel class IIa bacteriocin, piscicocin CS526, from Surimi-associated Carnobacterium piscicola CS526. Appl. Environ. Microbiol 2005, 71, 554–557. [Google Scholar]
  191. Gursky, L.J.; Martin, N.I.; Derksen, D.J.; van Belkum, M.J.; Kaur, K.; Vederas, J.C.; Stiles, M.E.; McMullen, L.M. Production of piscicolin 126 by Carnobacterium maltaromaticum UAL26 is controlled by temperature and induction peptide concentration. Arch. Microbiol 2006, 186, 317–325. [Google Scholar]
  192. Jack, R.W.; Wan, J.; Gordon, J.; Harmark, K.; Davidson, B.E.; Hillier, A.J.; Wettenhall, R.E.; Hickey, M.W.; Coventry, M.J. Characterization of the chemical and antimicrobial properties of piscicolin 126, a bacteriocin produced by Carnobacterium piscicola JG126. Appl. Environ. Microbiol 1996, 62, 2897–2903. [Google Scholar]
  193. Bhugaloo-Vial, P.; Dousset, X.; Metivier, A.; Sorokine, O.; Anglade, P.; Boyaval, P.; Marion, D. Purification and amino acid sequences of piscicocins V1a and V1b, two class IIa bacteriocins secreted by Carnobacterium piscicola V1 that display significantly different levels of specific inhibitory activity. Appl. Environ. Microbiol 1996, 62, 4410–4416. [Google Scholar]
  194. Hyronimus, B.; Le Marrec, C.; Urdaci, M.C. Coagulin, a bacteriocin-like inhibitory substance produced by Bacillus coagulans I4. J. Appl. Microbiol 1998, 85, 42–50. [Google Scholar]
  195. Motlagh, A.; Bukhtiyarova, M.; Ray, B. Complete nucleotide sequence of pSMB 74, a plasmid encoding the production of pediocin AcH in Pediococcus acidilactici. Lett. Appl. Microbiol 1994, 18, 305–312. [Google Scholar]
  196. Nieto, L.J.C.; Meyer, J.N.; Sletten, K.; Peláz, C.; Nes, I.F. Purification and amino acid sequence of a bacteriocin produced by Pediococcus acidilactici. J. Gen. Microbiol 1992, 138, 1985–1990. [Google Scholar]
  197. Marugg, J.D.; Gonzalez, C.F.; Kunka, B.S.; Ledeboer, A.M.; Pucci, M.J.; Toonen, M.Y.; Walker, S.A.; Zoetmulder, L.C.M.; Vandenbergh, P.A. Cloning, expression, and nucleotide-sequence of genes involved in production of pediocin PA-1, a bacteriocin from Pediococcus acidilactici PAC1.0. Appl. Environ. Microbiol 1992, 58, 2360–2367. [Google Scholar]
  198. Kim, I.K.; Kim, M.K.; Kim, J.Y.; Yim, H.S.; Cha, S.S.; Kang, S.O. High resolution crystal structure of PedB: A structural basis for the classification of pediocin-like immunity proteins. BMC Struct. Biol 2007, 7, 35. [Google Scholar]
  199. Rihakova, J.; Petit, V.W.; Demnerova, K.; Prévost, H.; Rebuffat, S.; Drider, D. Insights into structure-activity relationships in the c-terminal region of divercin V41, a class IIa bacteriocin with high-level antilisterial activity. Appl. Environ. Microbiol 2009, 75, 1811–1819. [Google Scholar]
  200. Tahiri, I.; Desbiens, M.; Benech, R.; Kheadr, E.; Lacroix, C.; Thibault, S.; Ouellet, D.; Fliss, I. Purification, characterization and amino acid sequencing of divergicin M35: a novel class IIa bacteriocin produced by Carnobacterium divergens M35. Int. J. Food Microbiol 2004, 97, 123–136. [Google Scholar]
  201. Ennahar, S.; Asou, Y.; Zendo, T.; Sonomoto, K.; Ishizaki, A. Biochemical and genetic evidence for production of enterocins A and B by Enterococcus faecium WHE 81. Int. J. Food Microbiol 2001, 70, 291–301. [Google Scholar]
  202. O’Keeffe, T.; Hill, C.; Ross, R.P. Characterization and heterologous expression of the genes encoding enterocin a production, immunity, and regulation in Enterococcus faecium DPC1146. Appl. Environ. Microbiol 1999, 65, 1506–1515. [Google Scholar]
  203. Morovský, M.; Pristas, P.; Javorský, P.; Nes, I.F.; Holo, H. Isolation and characterization of enterocin BC25 and occurrence of the entA gene among ruminal gram-positive cocci. Microbiol. Res 2001, 156, 133–138. [Google Scholar]
  204. Todokoro, D.; Tomita, H.; Inoue, T.; Ike, Y. Genetic analysis of bacteriocin 43 of vancomycin-resistant Enterococcus faecium. Appl. Environ. Microbiol 2006, 72, 6955–6964. [Google Scholar]
  205. Del Campo, R.; Tenorio, C.; Jiménez-díaz, R.; Rubio, C.; Gómez-Lui, R.; Baquero, F.; Torres, C. Bacteriocin production in vancomycin-resistant and vancomycin-susceptible Enterococcus isolates of different origins. Antimicrob. Agents Chemother 2001, 45, 905–912. [Google Scholar]
  206. De Kwaadsteniet, M.; Fraser, T.; van Reenen, C.A.; Dicks, L.M. Bacteriocin T8, a novel class IIa sec-dependent bacteriocin produced by Enterococcus faecium T8, isolated from vaginal secretions of children infected with human immunodeficiency virus. Appl. Environ. Microbiol 2006, 72, 4761–4766. [Google Scholar]
  207. Eguchi, T.; Kaminaka, K.; Shima, J.; Kawamoto, S.; Mori, K.; Choi, S.H.; Doi, K.; Ohmomo, S.; Ogata, S. Isolation and characterization of enterocin SE-K4 produced by thermophilic enterococci, Enterococcus faecalis K-4. Biosc. Biotechnol. Biochem 2001, 65, 247–253. [Google Scholar]
  208. Herbin, S.; Mathieu, F.; Brule, F.; Branlant, C.; Lefebvre, G.; Lebrihi, A. Characteristics and genetic determinants of bacteriocin activities produced by Carnobacterium piscicola CP5 isolated from cheese. Curr. Microbiol 1997, 35, 319–326. [Google Scholar]
  209. Hastings, J.W.; Sailer, M.; Johnson, K.; Roy, K.L.; Vederas, J.C.; Stiles, M.E. Characterization of leucocin A-UAL 187 and cloning of the bacteriocin gene from Leuconostoc gelidum. J. Bacteriol 1991, 173, 7491–7500. [Google Scholar]
  210. Van Reenen, C.A.; Chikindas, M.L.; van Zyl, W.H.; Dicks, L.M. Characterization and heterologous expression of a class IIa bacteriocin, plantaricin 423 from Lactobacillus plantarum 423, in Saccharomyces cerevisiae. Int. J. Food Microbiol 2003, 81, 29–40. [Google Scholar]
  211. Van Reenen, C.A.; van Zyl, W.H.; Dicks, L.M. Expression of the immunity protein of plantaricin 423, produced by Lactobacillus plantarum 423, and analysis of the plasmid encoding the bacteriocin. Appl. Environ. Microbiol 2006, 72, 7644–7651. [Google Scholar]
  212. Maré, L.; Wolfaardt, G.M.; Dicks, L.M.T. Adhesion of Lactobacillus plantarum 423 and Lactobacillus salivarius 241 to the intestinal tract of piglets, as recorded with fluorescent in situ hybridization (FISH), and production of plantaricin 423 by cells colonized to the ileum. J. Appl. Microbiol 2006, 100, 838–845. [Google Scholar]
  213. Atrih, A.; Rekhif, N.; Michel, M.; Lefebvre, G. Detection of bacteriocins produced by Lactobacillus plantarum isolated from different foods. Microbiology 1993, 75, 117–123. [Google Scholar]
  214. Atrih, A.; Rekhif, N.; Moir, A.J.; Lebrihi, A.; Lefebvre, G. Mode of action, purification and amino acid sequence of plantaricin C19, an anti-Listeria bacteriocin produced by Lactobacillus plantarum C19. Int. J. Food Microbiol 2001, 68, 93–104. [Google Scholar]
  215. Holck, A.; Axelsson, L.; Birkeland, S.E.; Aukrust, T.; Blom, H. Purification and amino acid sequence of sakacin A, a bacteriocin from Lactobacillus sake Lb706. J. Gen. Microbiol 1992, 138, 2715–2720. [Google Scholar]
  216. Axelsson, L.; Holck, A.; Birkeland, S.E.; Aukrust, T.; Blom, H. Cloning and nucleotide sequence of a gene from Lactobacillus sake Lb706 necessary for sakacin A production and immunity. Appl. Environ. Microbiol 1993, 59, 2868–2875. [Google Scholar]
  217. Axelsson, L.; Holck, A. The genes involved in production of and immunity to sakacin A, a bacteriocin from Lactobacillus sake Lb706. J. Bacteriol 1995, 177, 2125–2137. [Google Scholar]
Figure 1. Multiple sequence alignment of class IIa bacteriocins.
Figure 1. Multiple sequence alignment of class IIa bacteriocins.
Ijms 13 16668f1 1024
Figure 2. Organization of the gene clusters of class IIa bacteriocins. The figure was involved in production of avicin A in Enterococcus avium XA83 (avc, GenBank ID: FJ851402.1); bacteriocin MC4-1 in Enterococcus faecalis MC4 (bac, GenBank ID: EU047916.1); carnobacteriocin B2 in Carnobacterium maltaromaticum LV17B (cbn, GenBank ID: L47121.1); coagulin in Bacillus coagulans I4 (coa, GenBank ID: AF300457.1); divercin V41 in Carnobacterium divergens V41 (dvn, GenBank ID: AJ224003.1); enterocin A in Leuconostoc gelidum UAL 187 (ent, GenBank ID: AF099088); enterocin P in Enterococcus faecium P13 (ent, GenBank ID: AF005726.1); leucocin A in Leuconostoc gelidum UAL 187 (lca, GenBank ID: L40491.1); mesentericin 52A in Leuconostoc mesenteroides subsp. mesenteroides FR52 (mes, GenBank ID: AY286003.1); mundticin KS in Enterococcus mundtii NFRI 7393/AT06 (mun, GenBank ID: AB066267); mundticin L in E. mundtii CUGF08 (mun, GenBank ID: FJ899708.1); pediocin PA-1 in E. faecium Acr4 (pap, GenBank ID: HQ876214.1); penocin A in Pediococcus pentosaceus ATCC 25745 (pen, GenBank ID: NC_008525.1); piscicolin 126 in Carnobacterium piscicola JG126 (pis, GenBank ID: AF275938.1); plantaricin 423 in Lactobacillus plantarum 423 (pla, GenBank ID: AF304384); sakacin A in Lactobacillus sakei Lb706 (sap, GenBank ID: Z46867.1); sakacin G in Lactobacillus sakei CWBI-B1365 (skg, GenBank ID: EU570253.1) ; sakacin P in Lactobacillus sakei LTH673 (spp, GenBank ID: AF002276.1); sakacin X in L. sakei 5 (sak, GenBank ID: AAP44569.1); ubericin A in Streptococcus uberis E (uba, GenBank IDs: EF203953.1 and EF203954.1). Open reading frames (ORFs) encoding the related proteins are marked with the different color. The number of amino acid residues within each encoded protein is shown below the corresponding ORF.
Figure 2. Organization of the gene clusters of class IIa bacteriocins. The figure was involved in production of avicin A in Enterococcus avium XA83 (avc, GenBank ID: FJ851402.1); bacteriocin MC4-1 in Enterococcus faecalis MC4 (bac, GenBank ID: EU047916.1); carnobacteriocin B2 in Carnobacterium maltaromaticum LV17B (cbn, GenBank ID: L47121.1); coagulin in Bacillus coagulans I4 (coa, GenBank ID: AF300457.1); divercin V41 in Carnobacterium divergens V41 (dvn, GenBank ID: AJ224003.1); enterocin A in Leuconostoc gelidum UAL 187 (ent, GenBank ID: AF099088); enterocin P in Enterococcus faecium P13 (ent, GenBank ID: AF005726.1); leucocin A in Leuconostoc gelidum UAL 187 (lca, GenBank ID: L40491.1); mesentericin 52A in Leuconostoc mesenteroides subsp. mesenteroides FR52 (mes, GenBank ID: AY286003.1); mundticin KS in Enterococcus mundtii NFRI 7393/AT06 (mun, GenBank ID: AB066267); mundticin L in E. mundtii CUGF08 (mun, GenBank ID: FJ899708.1); pediocin PA-1 in E. faecium Acr4 (pap, GenBank ID: HQ876214.1); penocin A in Pediococcus pentosaceus ATCC 25745 (pen, GenBank ID: NC_008525.1); piscicolin 126 in Carnobacterium piscicola JG126 (pis, GenBank ID: AF275938.1); plantaricin 423 in Lactobacillus plantarum 423 (pla, GenBank ID: AF304384); sakacin A in Lactobacillus sakei Lb706 (sap, GenBank ID: Z46867.1); sakacin G in Lactobacillus sakei CWBI-B1365 (skg, GenBank ID: EU570253.1) ; sakacin P in Lactobacillus sakei LTH673 (spp, GenBank ID: AF002276.1); sakacin X in L. sakei 5 (sak, GenBank ID: AAP44569.1); ubericin A in Streptococcus uberis E (uba, GenBank IDs: EF203953.1 and EF203954.1). Open reading frames (ORFs) encoding the related proteins are marked with the different color. The number of amino acid residues within each encoded protein is shown below the corresponding ORF.
Ijms 13 16668f2a 1024Ijms 13 16668f2b 1024
Int. J. Mol. Sci. EISSN 1422-0067 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert
Back to Top