DIVERSITY OF BACILLUS SPECIES AND THEIR ANTIMICROBIAL COMPOUNDS INVOLVED IN ALKALINE-FERMENTATION OF INDIGENOUS FOOD CONDIMENTS USED IN AFRICA

Yérobessor Dabire 1,2 , Marius K. Somda 1 , Jerry Ugwuanyi 2 , Lewis I. Ezeogu 2 and Alfred S. Traore 1 . 1. Laboratory of Biotechnology in Food and Nutritional Sciences, Research Center in Biological Food and Nutritional Sciences, Department of Biochemistry and Microbiology, Research and Training Unit in Life and Earth Sciences, University Ouaga I, Prof. Joseph KI-ZERBO, 03 B.P 7031 Ougadougou 03, Burkina Faso. 2. Department of Microbiology, Faculty of Biological Sciences, University of Nigeria Nsukka, 410001, Enugu state, Nigeria. ...................................................................................................................... Manuscript Info Abstract ......................... ........................................................................ Manuscript History Received: 04 October 2018 Final Accepted: 06 November 2018 Published: December 2018

The indigenous food condiments, produced by alkaline fermentation of various African plant products, are widely used as food seasonings by most African people. Many strains of Bacillus genus are recognized as dominant microorganisms responsible of bioconversion of diverse plant-based seeds for the production of African alkaline-fermented food condiments. The involved Bacillus strains are known to produce a wide arsenal of useful antimicrobial compounds, particular polypeptides, lipopeptides and bacteriocins that exert broader spectra activities against Gram-negative and Gram-positive bacteria and fungi implicated in food toxicity or spoilage and ultimately human pathogenicity. Lipopeptides and bacteriocins present diverse biochemical structures with different mode and mechanism of action linked to their genetic and biosynthesis pathway. The molecular biology methods currently use in microbiological research allowed more reliable identification of antimicrobial polypeptides-producing Bacillus strains from these foods generating sufficient knowledge which potentiated the selection of starters cultures. The starters cultures and their antimicrobial peptides know a growing interest for effectiveness and best applications in many life domains. In this review, current knowledge about the main Bacillus species involved in African alkaline-fermented food condiments processes, mode and mechanism of action, genetic and biosynthesis pathway, and food applications of the antimicrobial peptides produced by these Bacillus strains are discussed.

ISSN: 2320-5407
Int. J. Adv. Res. 6 (12), 331-355 333 great growing interest to their effectiveness applications in food, biomedial and therapy, livestook and one others (Abriouel et al., 2011). In this paper, we are discussed on the mains Bacillus spp. involve in African indigenous alkaline-fermented food condiments production, the most known of their antimicrobial compounds, the mode and mechanism of action against foodborne or pathogen agents, the biosynthesis, and the potential applications of these antimicrobial compounds.

Diversity of Bacillus species from African indigenous alkaline-fermented food condiments
Alkaline food condiments are popular among most African countries. In addition to their important part in the diet (Parkouda et al., 2009), these food condiments play an economic, social and cultural role among African indigenous communities.
Several studies have focused on the microbiology diversity of alkaline fermented food condiments widely consumed by West African population. These studies generally reported Bacillus species as predominant microorganisms involved in the fermentation process of plant based-seeds for alkaline-fermented foods production (Parkouda et al., 2009). According to Omafuvbe et al. (2000), the alkaline fermentation is a process during which the pH of the substrate increases to alkaline values as high as pH 9.
Dawadawa, produced from the seeds of African locus beans (Parki biglobosa), is very popular in West Africa and plays an important role in many diets (Terlabie et al., 2006, Savadogo et al., 2011. Dawadawa, common name in Nigeria and Ghana, is also known under different local names in west Africa such as iru in Nigeria (Sanni et al., 2000), soumbala in Burkina Faso (Savadogo et al., 2011), netetu in Senegal (Ndir et al., 1994), afitin, iru and sonru in Benin (Azokpota et al., 2006) and Kinda in Sierra Leone. The isolation and identification of microorganisms in dawadawa from different countries of West Africa have recorded Bacillus species as the main microorganisms with the predominance of with Bacillus subtilis (Parkouda et al., 2009;Savadogo et al., 2011). B. subtilis, B. licheniformis and B. pumilus have mainly been found in dawadawa with predominance of B. subtilis (Odunfa and Oyewole, 1986;Terlabie et al., 2006). Ndir et al. (1994) reported Bacillus species as dominant microorganism in Senegalese netetu. Sarkar et al. (2002) recorded the predominance of B. subtilis in soumbala of Burkina Faso. Ouoba et al. (2004) also confirmed the predominance of B. subtilis and even B. pumilus in soumbala, and then consider the long cooking period during soumbala production as main key step for the selection of heat-resistant spore-forming of Bacillus species (Ouoba et al., 2007b). Beninese afitin, iru and sonru, have found to hold Bacillus spp. (Azokpota et al., 2006). Several Nigerian fermented food condiments are used as dishes flavor and/or a low-cost protein source (Sanni et al., 2002). Among others, we can cite, ugba from African oil bean (Pentaclethra macrophylla Bentham) seeds (Nurudeen et al., 2016); aisa from Albizia saman (Jacq.) F. Mull seeds (Ogunshe et al., 2006); okpehe, kpaye, or okpiye from mesquite (Prosopis Africana) seeds (Oguntoyinbo et al., 2007). Ogiri from melon (Citrullus vulgaris) seeds (Omafuvbe et al., 2004); and owoh from cotton (Gossypium hirsutum L.) seeds (Sanni and Ogbonna, 1991) are also listed in this group. All these food condiments have been investigated for their mean desirable microorganisms. The main fermenting microorganisms involved in the fermentation process of ugba were the proteolytic Bacillus species identified as B. subtilis (which is the most predominant), B. licheniformis, B. megaterium, B. macerans, and B. circulans (Nurudeen et al., 2016). Aisa has been found to harbor various Bacillus species as main microorganisms (Ogunshe et al., 2006). B. subtilis, B. licheniformis, B. megaterium, and B. pumilus were found as the main bacteria responsible for okpehe production (Ogunshe et al., 2007). B. pumilus, B. licheniformis, and B. subtilis are responsible of ground melon seeds fermentation for Ogiri production (Abaelu et al., 1990). Sanni and Ogbonna, (1991) reported the same species as dominant microorganisms in fermented cotton seeds, owoh.
Bikalga also known as dawadawa-botso in Niger, datou in Mali, furundu in Sudan and muja in Cameroon, is one of the most popular condiments in Burkina Faso used to flavor many dishes (Compaoré et al., 2013a). It is produced from alkaline fermentation of Hibiscus sabdariffa L. commonly known as Roselle or sorrel (Ouoba et al., 2008 (Compaoré et al., 2013a, b, c).

Antimicrobial peptides produced by Bacillus strains from alkaline -fermented food condiments
Bacillus strains produced at least two dozen different antimicrobial compounds that allow them to compete with other microorganisms in the same environment (Stein, 2005). Bacillus subtilis was found to devote an important portion of its genome (average 4 to 5%) for genes implicated in the production of antimicrobial compounds (Stein, 2005). Predominant B. subtilis group species isolated from African indigenous alkaline-fermented food condiments have been demonstrated to produce antimicrobial compounds potent against foodborne pathogenic bacteria and fungi (Ouoba et al., 2007b;Savadogo et al., 2011;Compaoré et al., 2013a, b, c). The most important of these antimicrobial compounds are the peptides group including lipopeptides (iturins, fengycins, and surfactins) and bacteriocins and/or Bacteriocins Like-Inhibitrice Substances (BLIS) that exhibit often a broad specific spectrum of antimicrobial activity (Savadogo et al., 2011;Compaoré et al., 2013a, b;Taalé et al., 20015). Using Polymerase Chain Reaction and Matrix Assisted Laser Desorption/ Ionization-Mass Spectrometry methods, Savadogo et al. (2011) identified surfactin from soumbala isolated Bacillus subtilis (S6, S21) strains. The ultra-high-performance liquid chromatography-time of flight mass spectrometry analysis allowed Compaore et al. (2013a, b) to identify surfactin and fengycin as main antimicrobial substances produced by B. subtilis ssp. subtilis H4 and B. amyloliquefaciens ssp. Plantarum strains isolated from Bikalga. Taalé et al. (2015) recorded BLIS production by Bacillus spp. strains Other antimicrobial compounds include polyketides (bacillaene, difficidin, and macrolactin), amino sugars and phospholipids (Stein, 2005).

Classification of antimicrobial peptides produces by Bacillus species
According to the biosynthesis pathways, the main antimicrobial peptides produced by Bacillus can be grouped in two dominant classes (Marx et al., 2001): the first group includes the non-ribosomally synthesized peptides, whereas the second comprises ribosomally synthesized peptides both presenting very diverse structures (Tapi et al., 2010).

Iturin family
With a molecular mass of ∼1.1 kDa, iturin is the smaller product among the 3 types of lipopeptides. The iturin family, including the related lipopeptides iturin A, C, D and E isoforms, bacillomycin D, F and L and mycosubtilin (Peypoux et al., 1986). All these compounds contain a cyclic heptapeptide composed of 7 amino acids interlinked or acylated with ß-amino fatty acids chain that can vary from C-14 to C-17 carbon atoms (Savadogo et al., 2011). Cyclic structure of lipopeptide iturin, consisted of seven amino acid residues attached to a 14-carbon chain, indicates its amphiphilic nature. Three D-amino acids (Tyr, Asn, and Asn) and four L-amino acids (Pro, Ser, Asn, and Gln) are the amino acids involved in this structure. (b) The cyclic heptapeptide surfactin contains both hydrophobic and hydrophilic amino acids. The structure containing amino acids: two D-amino acids (Leu, Leu) and five L-amino acids (Val, Asp, Leu, Glu, and Leu), indicates its amphipathic nature. (c) Primary cyclic structure of fengycin A. Structure containing peptide chain of ten amino acids and a -hydroxyfatty acid chain that can vary according to fengycin isomer from C-14 to C-17 carbons. In the structure, the amino acids are six L-amino acids a c b 336 (Glu, Glu, Pro, Gln, Tyr, and Ile) and four D-amino acids (Tyr, Orn, and Thr, Ala). Source: Khem and Shamsher, (2015). Iturin A comprises two major parts: a heptapeptide part and a hydrophobic tail of 11 to12 carbons ( Fig. 1(a)). This structure clearly shows an amphiphilic character of these compounds that targeted on the cellular membranes as the most probable site of their action (Aranda et al., 2005). Such molecules are of great interest because of their biological and physicochemical properties exploitable in food, oil, industry, etc… Source: Stein, 2005.

Surfactin family
The surfactin family contains a cyclic heptapeptide, with amino acids L or D, that form a lactone bridge with ßhydroxy (ß-OH) fatty acid. The ß-hydroxy fatty acid carbon chain length of C-13 to C-18 with amino acids sequence completely different from iturins (Magnet-Dana et al., 1992). Surfactin (∼1.36 kDa) is an amphipathic cyclic lipoheptapeptide of Glu-Leu-Leu-Val-Asp-Leu-Leu (ELLVDLL) containing the chiral sequence LLDLLDL interlinked with -hydroxy fatty acid chain consists of 12 to 16 carbon atoms to form a cyclic lactone ring structure ( Fig. 1(b)) (Seydlová et al., 2011). Different types of surfactin can be obtained according to the order of amino acids and the size of lipid portion, (Korenblum et al., 2012). Surfactin molecule contains hydrophobic amino acids located at positions 2, 3, 4, 6 and 7, and Glu and Asp residues located at positions 1 and 5, respectively. Surfactin isoforms usually coexist in the cell as a mixture of several peptidic variants with a different aliphatic chain length (Savadogo et al., 2011). The pattern of amino acids and -hydroxy fatty acids in the surfactin molecule depends both on producer bacterial strain and type of culture conditions (Seydlová et al., 2011). An intramolecular hydrogen bond can form the -turn, whereas the -sheet may depend on the same bond (Zou et al., 2010).

Fengycin family
Fengycin family includes cyclic lipodecapeptides formed by lactonization (Pathak et al., 2012). Fengycin structure consists of a saturated or unsaturated β-hydroxy fatty acid chain linked to the N-terminus of a cyclic decapeptide (Figure 1(c)) (Akpa et al., 2001). In the structure 8 amino acids (Tyr, Thr, Glu, Ala, Pro, Gln, Tyr, and Ile) of it decapeptide chain portion are involved in the formation of a peptide ring via lactone linkage between the side-chain phenolic-OH group of Tyr3 and C-terminal-COOH group of Ile10 (Pathak et al., 2012). Members of Fengycin family present heterogeneity both at the 6 th position in peptide moiety, and in -hydroxy fatty acid chain length that ranges from C-14 to C-17 carbon atoms and allows to obtain different homologous compounds and isomers (Kim et al., 2004). According to the variation of single amino acid at the 6 th position in peptide ring, fengycins have been classified in two classes: Fengycin A and fengycin B. Fengycin A contains Ala at position 6 and harbor unusual amino acids such as ornithine, while Val substituted Ala in case of fengycin B which isoforms harbor an allothreonine (Wang et al., 2004).

Bacteriocins and Bacteriocin-Like Inhibitory Substances (BLIS) Different classifications of Bacteriocins
Bacillus species produced bacteriocins and BLIS considered as the second most important after Lactic Acid Bacteria (LAB) bacteriocins. The main classification scheme for bacteriocins currently available is that of the LAB bacteriocins (Cotter et al., 2013). The classification scheme of LAB bacteriocins, firstly established by Klaenhammer (1993), has known subsequently many adaptations or reclassifications proposed by several authors taking account the chemical structure, the presence of modified amino acids, the molecular mass, the heat stability, the enzymatic sensitivity, the biosynthesis mechanism, and the mode of action of these bacteriocins (Arnison et al., 2013). According to above, Alvarez-Sieiro et al. (2016) proposed three major classes of LAB bacteriocins: Class I harbors small post-translationally modified peptides, Class II contains unmodified bacteriocins, and Class III holds larger peptides (>10 kDa, thermo-labile). Some antimicrobial peptides produced by Bacillus species have been included in the classes of LAB bacteriocins (as previously described by Klaenhammer, 1993). A good number of Bacillus bacteriocins belong to the group of lantibiotics ) included in the class I of LAB bacteriocins classification scheme (Nes et al., 2007). Furthermore, several bacteriocins/BLIS produced by Bacillus species belong to class II of LAB bacteriocins including the class IIa-pediocin-like bacteriocins and the two-peptide bacteriocins (class IIb) (Nes et al., 2007). However, some Bacillus lantibiotics described like halodurancin, lichenicidin, sublancin 168 or paenibacillin do not fit clearly in any of the classes of lantibiotics described above (Abriouel et al., 2011).

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Given the new elucidation of the structure, mode of export and mechanism of action of different described bacteriocins fall in various groups of bacteria, a unifying classification scheme of all bacteriocins still quite very difficult to establish (Makhloufi, 2011). So that, Abriouel et al. (2011) have proposed a separated classification scheme of Bacillus bacteriocins independently to that of LAB bacteriocins. Hence, until to date, only their classification scheme for Bacillus bacteriocins is available. This fact is most probably due to the lack of information on many of these bacteriocin amino acids sequences, and genes cluster and the considerable diversity of the peptides/proteins produced by the bacilli. According to Abriouel et al. (2011), there are three classes of Bacillus bacteriocins: Class I includes the post-translational modified peptides, Class II harbors the small non-modified and linear peptides, and Class III regroups large proteins. Moreover, the antimicrobial polypeptides and large antimicrobial proteins, unclearly identify are described under the category of BLIS (Abriouel et al., 2011).

The different classes of Bacillus bacteriocins
All the different classes of bacteriocins produced by Bacillus isolates are summarized in the Table 1

Subclass I.1: The lantibiotics
This subclass (Table 1) includes type-A lantibiotics (2.1 to 3.5 kDa) that are modified by a dual-enzyme system generically referred to as LanBC. They consist of 21 to 38 amino acid residues, exhibit more linear secondary structure and target the Gram-positive bacteria through voltage-dependent pores formation into the cytoplasmic membrane (Abriouel et al., 2011). Type-A lantibiotics include subtilin, the paradigm lantibiotic extensively studied in terms of its protein structure and genetic determinants. Subtilin (Fig. 2a) is a 3.32 kDa pentacyclic lantibiotic that contains 32-amino acid cationic, stable to acid and heat treatment up to 121°C for 30-60 min, and inhibits a broad range of Gram-positive bacteria including other Bacillus species . It is structurally similar to nisin from Lactobacillus lactis (Gálvez et al., 2007), and to B. . The mature ericin S and A reveal highly similar properties even though their precursors show only 75% identity. Only four amino acid residues distinguish ericin S to subtilin (Fig. 2a). Ericin S contains a subtilin-like lanthionine-bridging pattern that makes its antimicrobial activity and physico-chemical properties similar to subtilin. Different ring organization and 16 amino acid substitutions are held by ericin A compared with ericin S (Fig. 2a). Ericin A differs to subtilin by two C-terminal rings from it lanthionine pattern (Stein et al., 2002a).

Subclass I.2: the other single-peptide lantibiotics
The subclass I.2 includes type-B lantibiotics mersacidin, sublancin 168 and paenibacillin ( Table 1). The B. subtilis strain HIL Y-85.54728 mersacidin is a tetracyclic peptide with 1824 Da molecular mass (Bierbaum et al., 1995). Mersacidin displays a more globular structure due to the formation of four intermolecular thioether bridges or rings (

Class II bacteriocins: Non-modified peptides
This class forms a heterogeneous group of heat-stable, membrane-active peptides smaller than 10 kDa, that possess only standard amino acids and do not undergo post-translational modifications (Fernandez, 2014). They are subdivided in three subclasses: subclass II.1 named pediocin-like peptides, subclass II.2 or thuricin-like peptides, and subclass II.3 or linear peptides.

Subclass II.1: The Pediocin-like peptides
Bacillus Pediocin-Like peptides possess a disulfide bridge essential to the activity and a consensus motif YGNGVX1CX2K/NX3X4-C (X= any amino acid) at their N-terminal part. Their C-terminal portion is variable and can be both hydrophobic and hydrophilic (Feng et al., 2009). Coagulin (4612 Da) produced by B. coagulans I4, is an example of pediocin-like peptide, heat-stable and protease-sensitive, with anti-listeria activity (Table 1)

Mechanism of action of lipopeptides
Lipopeptides exert their antimicrobial activities by binding to the bacterial surface bilayer and alter the local lipid organizational linkages on negatively-charged fatty acids, and ultimately restructuring the lipid bilayer and thus preventing cellular processes.

Genetic organization and biosynthesis of Bacillus antimicrobial peptides Genetic organization and non-ribosomal biosynthesis of lipopeptides Genetic organization of lipopeptides.
Non-ribosomal biosynthesis uses in addition to 20 amino acids of the genetic code, other non-proteogenic molecules such as non-proteogenic amino acids (D forms, hydroxylated, methylated), fatty acids, sugars, lipids and hydrates of carbon from other biosynthetic pathways (Savadogo et al., 2011). It is for this reason that the term "monomers" is used to designate these different precursor molecules incorporated in non-ribosomal peptides (Tambadou, 2014).  Fengycins/plipastatins are also synthesized by NRPS encoded by an operon with five ORF ppsA-E (or fenA-E) (Fig.  3). The first three enzymes contain modules, the 4 th contains three modules and the last enzyme consists of one module (Ongena et al., 2008).
Iturin derivatives are synthesized by a PKS-NRPS hybrid complex unlikely to surfactin and fengycin (Stein, 2005). Iturin operon ranges from 38 to 40 kb in size and consists of four ORF, namely, fenF, mycA, mycB and mycC or ituD, ituA, ituB and ituC for mycosubtilin or iturin, respectively (Fig. 3) (Ongena et al., 2008). Three genes mycA (or ituA), mycB (or ituB) mycC (or ituC) encode for the NRPSs responsible for the incorporation of seven residues. The first one related to mycA (or ituA), the following four for mycB (or ituB) and the two last residues for mycC (or ituC). The structure of iturin A differs to that of mycosubtilin (in which the last amino acids are inverted) by an intragenic domain change in ituC and mycC. FenF (ituD) gene encodes a malonyl-CoA transacylase (MCT-domain). MycA also harbor PKSs related genes that are responsible for the final steps of the biosynthesis of the fatty acid chain (last elongation and β-amination) before it links up to the first amino acid of the peptidic moiety (Acyl-CoA ligase (AL-domain)) (Aron et al., 2005).

Non-ribosomal biosynthesis of lipopeptides.
Lipopeptides (iturins, surfactins and fengycins) are synthesized non-ribosomally via a multistep mechanism that involves the selection and condensation of amino-acid residues by modular megaenzymes NRPSs (Schwarzer et al., 2003). More than 300 different precursors are involved in the assemblage of these cyclic peptides, with possible branched structures containing a hydroxyl group, L-amino or D-amino acids, and furthermore modification by Nmethylation, acylation, glycosylation, or heterocyclic ring formation (Hancock and Chapple, 1999).

Genetic organization and ribosomal biosynthesis of bacteriocins. Genetic organization of bacteriocins.
The genes encode bacteriocins can be located on plasmids or on chromosomes (Kuipers et al., 2011). These genes are usually organized as an operon containing structural genes, immunity genes to protect the producer cell against its own synthesis, genes required for the transport and export of synthesized bacteriocin which are usually found on another locus and a gene encoding the leader peptide or prebacteriocin and genes for regulation (Jasniewski, 2008). A Quorum Sensing system mechanism regulates the bacteriocin production, and allows expression of certain genes according to the density of the bacterial population (Dortu and Thonart, 2009).

Genetic organization of class I bacteriocins Genetic organization of subclass I.1 bacteriocins
The genes involved in the biosynthesis of lantibiotics can be located on a transposon, on the chromosome or on a plasmid (Kuipers et al., 2011).
Subtilin is the paradigm lantibiotic extensively studied in terms of its protein structure and genetic determinants. Subtilin is structurally similar to nisin (Fig. 4a), and their respective biosynthetic gene clusters encode highly similar proteins (Siezen et al., 1996). The biosynthesis of subtilin is based on the expression of at least 10 genes from gene clusters spaBTCSIFEGRK (Fig. 4a) including the structural gene spaS for its precursor, genes spaB and spaC forproteins dehydratase (SpaB) and cyclase (SpaC), respectively, responsible for the post-translational modification of the presubtilin (Siezen et al., 1996). The cluster also includes gene spaT for the transporter SpaT, which associates with a membrane complex containing the enzymes SpaB and SpaC (Stein et al., 2003) and genes spaIFEG for immunity against the cognate bacteriocin.

Genetic organization of subclass I.2 bacteriocins
The lantibiotic mersacidin gene cluster is located in a region that corresponds to 3481 kbp on the chromosome of B. subtilis 168, (Fig. 4a) (Chatterjee et al., 1992). Its gene cluster contains 10 genes (spanning 12.3 kbp) including the structural gene mrsA, two genes (mrsM and mrsD) coding for precursor, one gene (mrsT) coding for a transporter with an associated protease domain and three genes (mrsF, mrsG, mrsE) coding for the group B, ABC Transporter involved in bacteriocin immunity (Altena et al., 2000). The cluster also includes three regulatory genes: two of them (mrsR2 and mrsK2) encode a two-component regulatory system apparently necessary for the transcription of the mrsFGE operon, and other gene mrsR1 encodes a protein involves in the regulation of mersacidin biosynthesis (Altena et al., 2000).
345 The lantibiotic sublancin 168 gene cluster is located on the chromosome of the producer B. subtilis 168 (Paik et al., 1998). The sublancin operon consists of five genes for it production and one immunity gene (Fig. 4a). The genes for the production include the structural gene sunA identified from the SPβ prophage region of the B. subtilis 168 chromosome, and four successive genes (sunT, bdbA, yolJ and bdbB) located downstream of sunA (Serizawa et al., 2005). The ORF sunT encodes a bifunctional ABC transporter that contains an ATP-binding cassette domain and a proteolytic domain (McAuliffe et al., 2001). The genes dbA and bdbB coed for thiol-disulfide oxidoreductases (Kouwen et al., 2007), while the function of yolJ is unknown. Upstream of the structural gene sunA is located the sublancin immunity gene sunI that encodes an immunity protein SunI, a membrane protein with a single N-terminal membrane-spanning domain (Dubois et al., 2009).

Genetic organization of subclass I.3 bacteriocins
Haloduracin from alkaliphile Bacillus halodurans C-125, consists of two post-translational peptides Halα/A1 and Halβ/A2 processed by the enzymes HalM1 and HalM2 responsible for their post-translational modification, respectively. The bioinformatics analysis has allowed to identify 11 potential genes spanning a 15 kbp region: the genes halA1 and A2 encode the haloduracin production, halM1, M2 code for modification, halt for transport and two sets of lanEFG homologs (halF1 to halE2) related to self-protection/immunity (Fig. 4a) (Fig. 4a). Indeed, lichenicidin gene cluster contains both two structural genes (licA1, A2), two genes (licM1, M2) coding for dedicated modification proteins of the LanM type, and a putative gene (licT) involved in the export and cleavage of the leader peptides to obtain the mature lichenicidin A and B subunits. The cluster also contains at least one set of genes of the lanEFG type dedicated in lantibiotic immunity (Begley et al., 2009). Some studies on the structural gene have recorded that the mature α-peptide Lica (Blia) (licA1, BLi04127) shared 40% similarity with mersacidin (Bierbaum et al., 1995) and 38% similarity with HalA1, while the mature β-peptide Licb(Blib) (licA2) showed 52% similarity to the haloduracin β-peptide HalA2 (McClerren et al., 2006).

Genetic organization of subclass I.4 bacteriocins
The operon coding for mature subtilosin A (Fig. 4a) consists of eight genes, sboA and albABCDEFG . The sboA gene is an operon that encodes the proteins involve in presubtilosin processing and subtilosin export and immunity. The albABCDEFG genes, transcribed from a promoter residing upstream of the sboA gene, are involved in the post-translational modification of presubtilosin. (Abriouel et al., 2011). The operon of sbo-alb, induced under anaerobic conditions, is controlled by two types of regulatory proteins: the transition state regulatory protein AbrB, and the two-component regulatory proteins ResD and ResE responsible for gene expression in response to limiting oxygen supply (Nakano et al., 2000).

Genetic organization of class II -bacteriocins
Generally, the biosynthesis of class II-bacteriocins mobilizes at least seven genes including an inducer gene, gene for protein kinase, regulatory gene, precursor gene (coding for the prebacteriocin), immunity gene, gene for the ABC transporter and a last gene encoding the transport accessory protein (Abriouel et al., 2011).

Genetic organization of subclass II.1 bacteriocins
The genes encoding the pediocin-like bacteriocins are organized into one or two operons and are located either on a plasmid, either on the chromosome (Belguesmia et al., 2011). The genetic determinants (coa operon) of coagulinis located on 14 kbp plasmid pI4 of its producer B. coagulans I4 (Le Marrec et al., 2000). These genetic determinants include an entire operon of four genes described for Pediococcus acidilactici pediocin PA-1/AcH (Fig. 4b), without the promoters for bacteriocin production. Downstream of the coa operon, a putative plasmid mobilization modules seem to be involved in plasmid transmission between bacteria (Le Marrec et al., 2000). The coagulin structural gene codes for a 44-amino-acid peptide similar to pediocin PA-1/ AcH, but possesses at position 41 a single C-terminal threonine residue, which is substituted by an asparagine (Asn41Thr) in a case of pediocin peptide. The coaB gene encodes bacteriocin immunity, while coaC and coaD are devoted to a secretion system mediated by an ABC transporter and its accessory protein, similar to those in P. acidilactici (Le Marrec et al., 2000).

Genetic organization of subclass II.2 bacteriocins
The B. thuringiensis thuricin 17 is encoded by three copies in tandem of the same structural gene (Fig. 4b). Each copy code for a 39-amino-acid precursor (Lee et al., 2009a). The three-gene copy is flanked upstream by secE gene encoding a protein translocase and nusG gene (a transcriptional anti-termination factor), and downstream by a sequence homologous to the albA gene of the subtilosin operon (Lee et al., 2009a).
The thurincin H (Fig. 4b). genetic determinants are held by bacterial chromosome. These genetic determinants consist of 10 ORFs including three structural genes (thnA1, A2 and A3) arranged in tandem repeats that are transcribed as a single transcript from a promoter upstream of the first structural gene as showed the RNA transcriptional analysis. The structural genes encode a 40-amino-acid prepeptide that evolves in a 31-amino-acid mature bacteriocin similar to thuricin 17 (Abriouel et al., 2011). The thurincin H structural genes are flanked downstream by a homologous gene to albA of thuricin 17 (thnB), followed by thnT and thnI genes, whichencode a putative ABC-transport protein and a hypothetical immunity protein, respectively (Lee et al., 2009b). The thurincin H cluster upstream region differs from that reported for thuricin 17. This region contains the putative transcriptional regulator thnR, thnD for ABC transporter system (ATP-binding protein) and thnE for permease protein. ThnP together with ThnB seem to be involved in bacteriocin processing. Indeed, the product of thnP showed homology to the epidermin leader-peptide processing by serine peptidase EpiP and to AlbB of B. subtilis (Abriouel et al., 2011).

Genetic organization of subclass II.3 bacteriocins
The members of subclass II.3 bacteriocins, cerein 7A and 7B are two non-synergistic bacteriocins produced simultaneously by Bacillus cereus Bc7, and different by their N-terminal amino acid sequences N-Gly-Trp-Gly-Asp-Val-Leu (7A) and N-Gly-Trp-Trp-Asn-Ser-Trp-Gly-Lys (7B) (Oscáriz et al., 2006). Only cerein 7B is characterized at molecular level. The cell DNA sequencing shown an ORF in the 416 contiguous nucleotides sequence that encoded a 74-amino acid protein containing a 18-amino acid leader sequence followed by 56 amino acids, corresponding to the mature cerein 7B, which structural gene promoter (Pribnow box) consisted of two 8 bp inverse repeats spanned by a 8 bp AT-rich region (Oscáriz et al. 2006).

Genetic organization of Class III bacteriocins
The large proteins megacin A-216 and megacin A-19213 produced by B. megaterium 216 and B. megaterium ATCC 19213, have their structural and immunity genes encoded on 211pBM309 (48 kb) and pBM113 (44 kb) 347 plasmid region in each Bacillus strains, respectively (Abriouel et al., 2011). The genetic determinants of megacin A-216 (Fig. 4b) encoded by a 5.494-bp plasmid region, include the structural gene megA, that codes for 293-aminoacid protein similar to proteins with phospholipase A2 activity, followed by an ORF encoding a 91-amino-acid protein dedicates to the producer self-protection. ORF 73 and gene encoding a 188-amino-acid protein similar to RNA polymerase factors, are at least required for the induction of megacin A-216 expression (Abriouel et al., 2011).

Ribosomal biosynthesis of bacteriocins of Bacillus
All bacteriocins are produced ribosomally in the cytoplasm of the producer cell as an inactive precursor called prebacteriocin recognized by the ABC transporter (ATP Binding Cassette transporter), which cleaves during or immediately after secretion to allow the bacteriocin to be active (Chen andHoover, 2003, Makhloufi, 2011. Furthermore, biosynthesis mechanism of class I bacteriocins involves variety of unusual amino acids (Abriouel et al., 2011). During maturation, the premature lantibiotics (precursor peptides) undergo intramolecular posttranslational modifications through the dehydration of serine and threonine residues and subsequent intramolecular addition of unusual thioether amino acids such as lanthionine and/or methyllanthionine residues to cysteine (Chatterjee et al., 2005). This addition results in the formation of (β-methyl) lanthionine thioether bridges, the characteristic structural elements for lantibiotics ), together with the proteolytic removal of leader peptides (Dischinger et al., 2009).

Application of antimicrobial peptides produced by Bacillus strains
Bacillus antimicrobial peptides, given to their safety, received more attention todays for several applications such as biomedical and therapeutic, pharmaceutical and food applications (Abriouel et al., 2011). Only food applications of lipopeptides and bacteriocins produce by Bacillus species are discussed and the other applications being beyond of the scope of this review.
To date, consumer demands for minimally processed foods or 'fresh foods' without chemical preservatives have stimulated research interest in the new bioconservation methods (Abriouel et al., 2011;Silva et al., 2018). The bioconservation of a food is to increase its shelf life and improve its safety by using microorganisms and/or their metabolites (Ross et al., 2002). Indeed, the screening of antimicrobial peptides for food applications requires the fulfillment of some extensive criteria (EFSA, 2008). Producer strains should be food grade (GRAS or QPS), exhibit a broad spectrum of inhibition, exert high specific activity, have health risks free, present beneficial effects (e.g. improve safety, quality, and flavor of foods), display heat and pH stability, and optimal solubility and stability for a particular food (Mills et al., 2011). Antimicrobial peptides-producing Bacillus strains or their peptides, which meet the above criteria, could be used as safe food biopreservatives even thought, their applications are rarely evaluated compared to LAB bacteriocins (e.g. nisin and pediocin PA1), which have the GRAS status (Simha et al., 2012).
In the food industry, lipopeptides are used as emulsifiers during processing of raw materials. The baking industry uses surfactins to maintain the texture, stability, and volume of fat, and also its emulsification ).
An increasing interest is also granted to Bacillus bacteriocins for food preservation since they are superior to LAB bacteriocins (Abriouel et al., 2011). Usually, the use of bacteriocins as food preservatives includes the main following approaches: inoculation of food with the bacteriocin-producing strain (starters culture or protective cultures), useof food previously fermented with a bacteriocin-producing strain as an ingredient in food processing, and the addition of purified or semi-purified bacteriocin as food additives, with the requirement of express authorization of their use. Unfortunately, there are not enough data on the possible applications of lipopeptides and bacteriocins produced by the various mains Bacillus strains isolated from African indigenous alkaline-fermented food condiments. However, application of lipopeptide-or bacteriocin-producing Bacillus strains in these food condiments and beverage substrates know new opportunities in food biopreservation. Hence, subtilisin-producing B. subtilis strain, an inhibitor of toxin-producing B. cereus, has been suggested as starter culture to improve the safety in the production of okpehe (Oguntoyinbo et al., 2010). BLIS-producing B. subtilis B7 and B15 strains has been proposed as starter cultures for soumbala production (Ouoba et al., 2008a). Strains B. subtilis subsp. subtilis H4 and B. amyloliquefaciens ssp. plantarum produced both lipopeptides (iturins, surfactins and fengycins) and bacteriocins 348 active against B. cereus, L. monocytogenes, M. luteus and S. aureus, and has been proposed as a useful starter cultures for safe production and biopreservation of Bikalga (Compaoré et al. (2013a, c). BLIS-producing B. subtilis, with a wide antimicrobial spectrum, has promising application in food biopreservation (Taalé et al., 20015).

Economic impact of antimicrobial peptides.
The sale values of food additives are knowing continuous growth rates about 2 to 3% annually, and especially for emulsifiers and hydrocolloids in terms of market increase (Freire et al., 2009). It is quite likely that lipopeptides and bacteriocins produced by Bacillus species, in the near future, will represent significant percentage of food additive in the market.
The bacteriocins are used in purified, semi-purified form or in the form of a concentrate obtained after fermentation of a food substrate (Makhloufi, 2011). It is difficult to condition the bacteriocins in purified form because their purification is expensive and it requires several steps (precipitation -chromatography on column -reverse phase HPLC), hence difficulties for production on an industrial scale. Adsorption technique is most used because of the cationic nature of bacteriocins. Several forms of conditioning of bacteriocins are to date proposed: adsorption of bacteriocins in silicone particles, encapsulation in liposomes or incorporation into different materials (calcium aliginate, cellulose, soy protein, polysaccharide films), and freeze-dried (Sutyak et al., 2008b). All these problems encountered in the production and packaging of bacteriocins explain their low rate of use as a food preservative or for other applications. Nevertheless, the bacteriocin trade has an annual growth rate of 2-3% and has reached a turnover of over

Implications antimicrobial peptides for food safety and shelf-life
Several studies have demonstrated that the desirable Bacillus strains from alkaline-fermented foods produce useful lipopeptides and bacteriocins that exert some broader spectra against spoilage and pathogenic microorgisms (Ouoba et al., 2008). Thereby, these antimicrobial peptides contribute to food safety and enhance it shelf-life (Compaoré et al., 2013b, c;Nah et al., 2015). However, efforts still have to be made to use desirable Bacillus strains and/or their antimicrobial peptides, either alone or in combination with the mild physicochemical treatments and low concentrations of traditional and natural chemical preservative, as an efficient way of extending shelf-life and food safety through the inhibition of spoilage and pathogenic bacteria without altering the nutritional value of food products in order to meet the consumer demands.

Conclusion and future perspectives
Bacillus species particularly the member of Bacillus subtilis group are the main bacteria involve in the plant-based seeds bioconversion for African alkaline-fermented food condiments production. The indigenous alkaline fermented condiments are being promoted in some countries due to their beneficial role in the diet, and there is increasing demand for such products in both rural and urban areas. Indeed, the ability of many Bacillus strains to produce lipopeptides and bacteriocins, mainly active against foodborne spoilage and pathogens, and human and/or animal pathogens, leads to suppose that the GRAS status could be further extended to some Bacillus species and/or their antimicrobial peptides for food productions in industrial level. Several studies shown that lipopeptides and bacteriocins represent a group of bioactive molecules with a diversity of structures offering a broad spectrum of physico-chemical properties that can lead to various industrial applications, meeting the needs of consumers and industries. Scientists and industrialists are increasingly interested in the biological activities of these molecules and their potential food applications as well in other fields. On this point, the scientific literature is regularly enriched with data relating to the potential antimicrobial peptides for applications in different life sector.

Conflict of Interests statements
The authors declare that there is no conflict of interest for this article.