Review Bioscience Microflora Vol. 21 (4), 217-223,2002 Antimicrobial Substances Including Produced by Lactic Acid NagendraP. SHAH1*and RajivDAVE2 SchoolofLife Sciences, Victoria University ofTechnology, POBox 14428, Melbourne CityMail Centre, Victoria 8001, Australia 2Currentaddress: Dairy Science Department , SouthDakota State University, Brookings, SD57007, USA Receivedforpublication, May9, 2002

Thisbrief overview highlights the antimicrobialsubstances including bacteriocins produced by , witha particularemphasis on bacteriocinproduced by probioticbacteria, such as Lactobacillusacidophilus and bifidobacteria. Keywords: antimicrobialsubstances; bacteriocins; Lactobacillus acidophilus; bifidobacteria; nisin

probiotic bacteria such as Lb. acidophilus and bifido- INTRODUCTION bacteria. A is defined as a proteinaceous compound ANTIMICROBIALSUBSTANCES PRODUCED produced by many different bacterial species that pos- BY DAIRY LACTIC ACID BACTERIA AND sess bactericidal action against other microorganisms, THEIR INHIBITORYSPECTRUM which are closely related to the producer organisms. A large number of bacterial species are reported to pro- Organic Acids duce bacteriocins including colicin from Escherichia Lactic acid bacteria while growing in milk produce coli, subtilin from Bacillus subtilis, pediocin by mainly lactic, formic, acetic, and propionic acids. These Pediococcus pentosaceus, and nisin from Lactococcus acids help lower the pH and create unfavourable envi- lactis subsp. lactis. The bacteriocins produced by spoil- ronment for other microorganisms. It has been demon- age and pathogenic group of microorganisms are not strated that bacteriostatic effect of lactic acid is pri- reviewed here, as they are precluded from food use due marily due to lactate anions rather than hydrogen ions. to the pathogenicity of producer organisms. In addi- The presence of lactic acid in fermented milk foods is tion to bacteriocins, lactic acid bacteria (LAB) produce shown to be inhibitory towards Staphylococcus and other antimicrobial substances such as lactic acid, Yersinia and bactericidal to Campylobacter and Sal- diacetyl, hydrogen peroxide, and bacteriocin-like in- monella. Lactic acid and/or propionic acid are effec- hibitory substances (5). LAB and the antimicrobial sub- tive against Campylobacter. Propionic acid produced stances produced by these bacteria have been used to by dairy propionibacteria has a broader inhibitory spec- produce safe and wholesome foods for centuries. The trum than lactic acid or acetic acid and inhibits moulds inhibitory activity of LAB leads to the creation of hos- and Gram-negative bacteria, but is ineffective against tile environment for food-borne pathogens and spoil- Gram-positive species. age organisms in foods.. The inhibition of pathogens and spoilage organisms is due to production of lactic Hydrogen Peroxide and other organic acids, hydrogen peroxide, diacetyl, Some strains of LAB including lactococci, lactoba- and bacteriocin, competition and nutrient depletion, and cilli, leuconostocs, and pediococci have the ability to altered redox potential. However, it is generally believed generate hydrogen peroxide during growth and lack of that the inhibitory activity is not a function of any one catalase by these bacteria causes its accumulation in compound, but it is a composite effect of several fac- growth media. Accumulation of hydrogen peroxide tors. This review will focus on the antimicrobial sub- occurs by the action of superoxide dismutase in most stances including bacteriocin produced by LAB, with lactic bacteria (particularly lactococci) or by manga- a particular emphasis on bacteriocins produced by nese ions present in high concentrations in the cyto- plasm of bacteria (particularly lactobacilli, leuconostocs *Corresponding author . Mailing address: School of Life Sciences, Victoria and pediococci). Lactococci produce sufficient hydro- University of Technology, PO Box 14428, Melbourne City Mail Centre, Victoria 8001, Australia. Phone: +61-3-9216-8289. Fax. +61-3-9216-8284. gen peroxide to be auto-inhibitory. Hydrogen peroxide E-mail: [email protected] has been reported to inhibit the growth of Staphylococ-

217 218 N.P. SHAH and R. DAVE cus aureus, E. coli, Salmonella typhimurium, Clostrid- so-called . Lantibiotics are small ribosomally ium perfringens, Pseudomonas sp. and other psy- synthesised polypeptides containing modified amino chrotrophs. However, the concentration of hydrogen acids such as lanthionine and 3-methyl-lanthionine. The peroxide produced by starters may not be sufficient to most prominent lantibiotic is nisin. Class I bacteriocin directly affect the microbial cells in foods. In raw milk, undergo post-translational modifications. hydrogen peroxide generated by LAB can react with The class II bacteriocins are small hydrophobic and endogenous thiocyanate, a reaction catalysed by lac- heat-stable peptides, which do not contain unusual toperoxidase, to form intermediary oxidation products amino acids such as lanthionine. To date, many bacte- inhibitory to microorganisms. riocins belonging to class II have been identified and characterised such as Lactacin F, Lactococcin A, B and Diacetyl M, Leucosin, Plantaricin A, Pediocin PA-1, and Sakacin Diacetyl (2,3-butanedione) is a metabolic end prod- P, Curvacin A, Enterocin A. Class II bacteriocin pep- uct of LAB synthesised from the intermediary metabo- tides do not undergo post-translational modifications lite, pyruvate and is primarily known as flavour com- and contain between 30 and 60 residues. Class II bac- pound in fermented dairy products. Certain species of teriocins have two sub-groupings: class IIa bacterio- LAB have the ability to synthesise diacetyl, which can cins are effective against and thus have poten- inhibit yeasts, and several Gram-positive and Gram- tial as antimicrobial agent in food and feed. The major- negative organisms. The concentration required for such ity of bacteriocins produced by L. acidophilus are heat inhibition ranged from 100 to 400 ƒÊg/ml, while lactic stable, low molecular mass, non-lanthiobiotic peptides, starters produce only 10 to 80ƒÊg/ml of diacetyl . Thus, which belong to class II. The production of bacterio- diacetyl may be a minor contributor to the broad-spec- cins of Carnobacterium piscicola LV 17, Lb. plantarum trum antagonism. Cl 1 and Lactobacillus sake Lb 706 is induced by pep- tides excreted by producer cells. The peptides making Bacteriocins up the bacteriocin do not contain lanthionine residues Lactic acid bacteria produce a wide variety of anti- and such type of bacteriocin is the most prevalent bac- microbial including , anti- teriocin produced by LAB. biotic-like substances, bacteriocins and bacteriocin-like Class III bacteriocins contain large heat labile pep- substances for the inhibition of food-borne pathogens tides (>30 kDa). A typical example is bacteriocin pro- and spoilage organisms. Among the like sub- duced by L. helveticus, helveticin J. Class IV bacterio- stances, nisin is well characterised. Bacteriocins are cins are complex bacteriocins formed by the associa- proteinaceous compounds that show antimicrobial 'ac- tion of bactericidal proteins with one or more other es- tivity against closely related species' (7). While the sential chemical moieties such as lipid and carbohy- definition holds true for majority of bacteriocins, it is drate moieties. now evident that bacteriocins may act beyond closely Most bacteriocins produced by LAB have narrow an- related species or those confined to the same ecologi- tibacterial spectrum, confined to species related to pro- cal niche. ducer organisms, whereas some bacteriocins are active Among the bacteriocins, nisin is the one most exten- against Listeria sp. are and other food-borne pathogenic sively characterised and it is widely used as a food pre- and spoilage organisms. Most bacteriocins are hydro- servative in several countries. Other commercial prod- phobic and hence can be bound by lipids and phospho- ucts used by food industry include MicrogardTM (a prod- lipids. uct containing several metabolites produced by fermen- The antimicrobial activity of bacteriocins is due to tation with Propionibacterium freudenreichii subsp . increases in the permeability of the cytoplasmic mem- shermanii), which is used in cottage cheese. brane of target cells causing the dissipation of the pro- ton motive force or disturbing membrane transport and CLASSES OF BACTERIOCINS AND thus inhibiting energy production and biosynthesis of MODE OF ACTION proteins. The mechanism of action of nisin involves Biochemical and genetic studies of bacteriocins pro- binding to the peptidoglycan layer, causing destabilisa- duced by lactic acid bacteria have now defined four tion of the membrane by the formation of pores, which major classes (4). Bacteriocins of LAB belong to ei- allow leakage of ions such as potassium and magne- ther class I or class II. Class I bacteriocins are mem- sium and dissipation of the proton motive force. Gram- brane-active and heat-stable peptides. They contain the negative organisms are resistant to hydrophobic mol- ANTIMICROBIAL SUBSTANCES INCLUDING BACTERIOCINS PRODUCED BY LACTIC ACID BACTERIA 219 ecules like nisin due to outer membrane permeability licic acid column from an active fraction in the acid barrier. Nisin can be either bacteriostatic or bactericidal. soluble fraction of cultures of Lb. acidophilus. Lacto- Nisin action against susceptible Gram-positive vegeta- cidin has a broad antibiotic spectrum against Gram- tive cells results from molecular damage to the cyto- negative and Gram-positive bacteria. plasmic membrane. Nisin causes rapid leakage of cel- Acidophilin, a low molecular-weight•E peptide, has been lular material, including ATP and various ions, and extracted from methanol and acetone extract of aci- complete loss of membrane potential. The effect on dophilus milk. Acidophilin is heat-stable under acidic spore forming bacteria is sporostatic. The bacteriocin conditions and exerts antibacterial action against lactococcin A specifically increases permeability of pathogenic bacteria in vitro. lactococcal cytoplasmic membranes in a voltage-inde- Acidolin has been isolated•E and purified on Sephadex pendent -mediated manner. The voltage inde- G25 from methanol extracts of acidophilus milk fer- pendent activity of lactococcin B is dependent on the mented with Lb. acidophilus 2181. It has a molecular reduced state of its only residue. Some bacte- weight of 200 Da. Acidolin is extremely heat-stable riocins including pediocin AcH, leucocin Lcml, sakacin and its solution is strongly acidic. It has a broad anti-

A and nisin adsorb to the producer cells. Reports also bacterial spectrum, particularly against spore-form- indicate that bacteriocins act as lethal particulates and ing bacteria. Acidolin is responsible for antibacterial can kill the producer organism termed as "quantal" kill- activity in vitro against various spoilage and patho- ing. genic type organisms. Further details of bacteriocins and their mode of ac- extract •Eof Lb. acidophilus ATCC 3205 produced tion can be found elsewhere (2, 8). a peptide of molecular weight of 3,500 Da. The bac- teriocin shows antibacterial activity against E. coli

Bacteriocins Produced by Lactobacilli and Bifidobac- over a wide range of pH. terium spp. Lactocin B has been •Eisolated and purified from the

Lactobacilli and bifidobacteria are becoming increas- active fraction of filtrate from cultures of Lb. acido- ingly popular as starters due to their probiotic effects. philus N2 by ion-exchange chromatography, ultrafil- The main reasons for their popularity are: (i) their habitat tration and gel filtration. Lactocin has a molecular in the gastrointestinal tract, (ii) their recognition as safe weight of 6,000-6,500 Da. Its antibacterial activity is bacteria, and (iii) their ability to control spoilage and restricted to lactobacilli. putrefying bacteria in the gut. The use of L. acidophi- Bacteriocins •Eof Lb. acidophilus AC1 and AR1 have lus and Bifidobacterium sp. in foods has drawn atten- been isolated and purified using ammonium sulphate tion of scientists to study the details of antimicrobial precipitation of filtrates from cultures of Lb. acido- systems of these bacteria, which are briefly summarised philus strains AC1 and AR1. The Lb. acidophilus AC1 below. bacteriocin has a molecular weight of 5,400 Da, whereas that of Lb. acidophilus AR1 5,200 Da. These

Bacteriocins of Lactobacilli bacteriocins show antibacterial activity against B. Among lactobacilli, Lb. acidophilus has been re- subtilis, S. aureus, E. coli and Salmonella sp. The Lb. garded as a good candidate for use as dietary adjunct. acidophilus AC 1 bacteriocin lost its activity when The chemical nature and structure of antibacterial sub- heated at 50•Ž for 20 min, but the Lb. acidophilus stances produced by Lb. acidophilus have been studied AR1 bacteriocin was stable when heated for 80 min extensively. at 50•Ž under acidic conditions. Both substances show Lb. acidophilus produces organic acids, hydrogen an optimum pH of 4.0-5.5 and both are inactivated peroxide and bacteriocins to suppress the multiplica- by treatment with trypsin. tion of pathogenic and putrefying bacteria. Lb. acido- Bacteriocin produced •E by Lb. acidophilus LAPT 1060 philus shows stronger antibacterial properties against isolated from infant faeces was found to inhibit six Gram-positive bacteria such as S. aureus and C. strains each of Lb. delbrueckii subsp. bulgaricus and perfringens than against Gram-negative bacteria such Lb. helveticus. The bacteriocin has been designated as S. typhimurium and E. coli. as 'acidophilucin A' and is sensitive to proteolytic

Several bacteriocins produced by Lb. acidophilus enzymes and heat (60•Ž for 10 min). Antibacterial have been reported as described below. substance produced by Lb. acidophilus LA-147 is in-

A bacteriocin referred to as •ELactocidin has been activated by heat (100•Ž for 15 min), protease and a- identified and purified by chromatography on a si- chymotrypsin treatments. 220 N.P. SHAH and R. DAVE

Table 1. Bacteriocin and bacteriocin-like inhibitory substances produced by lactobacilli (examples) .

Adapted from various sources.

・ Antimicrobial substances produced by Lb. acidophi- by Lb. acidophilus has been reported by Dave and lus, Lb. fermentum, and Lb. plantarum are effective Shah (1). against several strains of Lb. delbrueckii subsp. The bacteriocins of lactobacilli have been listed in bulgaricus, Lb. casei, Lb. helveticus, and Lb. jugurti. Table 1. The production of bacteriocin occurs in MRS broth, but very little bacteriocin is produced in milk or whey- Bacteriocin of Bifidobacteria based medium In general, bifidobacteria have higher antibacterial ・ Antimicrobial substance produced by Lb. helveticus activity compared to lactobacilli. However, most bifido- inhibits the growth of several strains of Lb. delbrueckii bacteria do not produce any antibacterial substances subsp. bulgaricus, several strains of Lb. acidophilus, other than lactic acid and acetic acid. Only few reports Lb. plantarum, and Lb. jugurti. The bacteriocin is are available on the nature of the anti-microbial activ- found to be active over a pH range of 3 to 10. Its ac- ity of bifidobacteria and studies on the chemical nature tivity against Lb. acidophilus is inactivated at 70•Ž . and structure of antibacterial substances produced by Inhibition of several spoilage and pathogenic type of bifidobacteria are still in infancy stage. microorganisms by the bacteria is considered pri- ・ Ofthe five strains of Bifidobacterium.bifidum (ATCC marily due to acid. 11863, 29591, and NCFB 1453, 1454 and 1455) ex- ・ Inhibition of Lb. acidophilus by antimicrobial sub- amined for production of bacteriocins in MRS broth stance produced by Lb. helveticus may be of concern with 0.05% cysteine, only one strain, Bif. bifidum as both species are used to manufacture AB (Acido- NCFB 1454, excreted a bacteriocin, designated as philus-Bifidus) yogurt and both organisms may be bifidocin B. The bacteriocin is active against Gram present in the same product. positive bacteria but not against Gram negative bac- ・ Inhibition of several strains of Lb. delbrueckii subsp. teria. The amino acid sequence indicated that the bac- bulgaricus, Lb. helveticus, Lb. jugurti and Lb. casei teriocin contained 36 amino acid residues. Bifidocin ANTIMICROBIAL SUBSTANCES INCLUDING BACTERIOCINS PRODUCED BY LACTIC ACID BACTERIA 221

Table 2. Bacteriocins produced by lactococci, leuconostocs and pediococci (examples).

Adapted from various sources.

B is active against some food-borne pathogens such active against Listeria and other food-borne pathogenic

as Listeria, Enterococcus, Bacillus, Lactobacillus, and spoilage organisms. Most bacteriocins are hydro-

Leuconostoc and Pediococcus sp. phobic and hence can be bound by fats and phospho- A bacteriocin named bifidin, from•E B. bifidum 1452, lipids. Nisin causes cellular death by affecting cyto-

has been purified. The substance is found to be heat plasmic membrane and proton motive force (3). stable at 100•Ž for 30 min. The main amino acids of Nisin was discovered in 1928 and has received com- bifidin are phenylalanine and glutamic acid. Bifidin mercial application in the food industry. The molecu- shows antibacterial activity against Micrococcus lar structure of nisin has been determined. Nisin has

flavus and S. aureus and the compound is active at shown to be 3,500 Da peptide consisting of unusual

pH 4.8-5.5. amino acids. It is made up of pentacyclic peptide of 34 amino acids residues containing the three unusual amino BACTERIOCIN PRODUCED BY LACTOCOCCUS, acids didehydroalanine (Dha) (residues 5 and 33), ƒÀ- LEUCONOSTOC, AND PEDIOCOCCUS methyldidehydroalanine (ƒÀ-MeDha) or didehydro-

Currently known bacteriocins produced by Lacto- butyrine (Dhb) (residue 2) and the thioether amino ac- coccus, Leuconostoc, and Pediococcus and their inhibi- ids lanthionine (Lan, Ala-S-Ala) (residues 3-7) and ƒÀ- tory spectrum are listed in Table 2. Of these, nisin and ethyllanthionine (ƒÀ-MeLan, Ala-S-Aba; mAba, pediocin have been best characterised and have found aminobutyric acid) (residues 8-11, 13-19, 23-26, and food applications. 25-28). Hence, the nisin molecule contains three a, ƒÀ- unsaturated amino acids and five intramolecular sulfur

Nisin rings formed by one lanthionine residue (ring A) and Among the bacteriocins produced by dairy LAB, ni- four ƒÀ-methyllanthionine residues (rings B, C, D, and sin is best characterised. Nisin is produced by Lacto- E). Because of these lanthionine moieties, nisin is coccus lactis subsp. lactis and has a broad spectrum. known as a lantibiotic.

Most bacteriocins produced by LAB have a narrow Nisin is permitted as a food additive in at least 47 antibacterial spectrum, whereas some bacteriocins are countries. Nisin has been given GRAS status in US in 222 N.P. SHAH and R. DAVE

processed cheese. Nisin has been successfully incor- Ped. acidilactici PAC 1.0 is stable to heating at 100•Ž, porated for preservation in several food commodities but the activity is reduced at 121•Ž. The amino acid including processed cheese, hard cheese, milk, yogurt, sequence of pediocin PA-1 and pediocin AcH has cottage cheese, bacon and smoked fish, but mainly in shown that the two bacteriocins are the same. the production of processed cheese. Both pediocins have been used in food systems to

Because nisin is heat-stable, it can be added before control the growth of Listeria monocytogenes. The heat processing or canning of foods. Since it is a poly- antilisterial activity of the bacteriocin is greater at higher peptide, any residues remaining in foods are digested. pH. Inhibition of growth of L. monocytogenes in frank- Further, it acts as an adjunct to heat processing and furters by pediocin has also been reported. A number supplementary heat inactivation can be achieved even of factors need to be considered for bacteriocidal effi- at a comparatively low temperature. All these proper- ciency of pediocin AcH against L. monocytogenes in- ties have made nisin a very useful food preservative. cluding pH, concentrations of sodium chloride, texture, Nisin can be either bacteriostatic or bacteriocidal, de- and lipid content. Pediocin AcH is also reported to con- pending on the target organism. It has a sporostatic ef- trol the growth of spoilage strain of Clostridium laramie. fect on bacterial spores. Growth of Bac. subtilis at 106 In general studies have shown that pediocin PA-1 and cells/g is completely inhibited by nisin producing pediocin AcH can be used as a biopreservative to con- Lactococcus lactis ATCC 19659. The mechanism of trol both spoilage and pathogenic Gram-positive bac- action of nisin against Sal. typhimurium and verotoxi- teria. genic strains of E. coli serotypes O157:H7, O104:H22 BACTERIOCIN LIKE INHIBITORY SUBSTANCES and O111:NM has been examined. During 14 days at

6-7•Ž, nisin produced a 1.5-2.0 log cycle reduction in A number of lactic acid bacteria produce bacterio- the counts of both groups of bacteria in peptone water. cin-like inhibitory substances (BLIS) (6). The best ex- Nisin has been shown to sensitise Bacillus sp. spores ample of BLIS is reuterin. Reuterin, produced by Lac- to heat and presence of residual nisin in the recovery tobacillus reuteri, a prominent member of the Lacto- medium prevents the outgrowth of survivors. bacillus population has been well characterised. Nisin has been used to control spore-forming bacte- Reuterin has a molecular mass of < 200, it is resistant ria including Cl. sporogenes, Cl. tyrobutyricum. and to protease activity, and therefore it does not fall into

Bacillus, which are frequently associated with the spoil- the definition of bacteriocin. Reuterin is a potent, broad- age of processed cheese spreads. Nisin is also effective spectrum BLIS effective against Gram-negative (e.g., against Staphylococcus, Listeria, Salmonella, Leu- Salmonella and Shigella), Gram-positive (e.g., conostoc, Pediococcus, Lactobacillus, and Micrococ- Clostridium and Listeria), yeasts, fungi and protozoa. cus. It is primarily inhibitory to Gram-positive bacte- Reuterin has been used as a natural preservative for ria, but when used together with EDTA, it effectively food and feed as well as for preservation of fish. inhibits Gram-negative bacteria. APPLICATIONS Bacteriocins formed by Lc. lactis strains ITAL 383 and CNRZ 150 are similar to nisin formed by L. lactis Antibacterial substances produced by LAB could be strain ATCC 11454. The bacteriocin is bactericidal and utilised for food processing to inhibit the growth of causes lysis of Listeria innocua strain LIN 11. pathogenic and spoilage type organisms. Because bac- teriocins are natural products, there is a tremendous

Pediocin interest in their use as a novel means to ensure the safety Bacteriocin production by Ped. pentosaceus (previ- of minimally processed refrigerated foods. The poten- ously known as Ped. cerevisiae) and Ped. acidilactici tial uses of bacteriocins as food preservative include has been reported. These bacteriocins inhibited growth meat, fish, dairy, cereal, fruits and vegetables. of selected strains of Gram-positive bacteria including Today, only nisin, a well-characterised bacteriocin Pediococcus, Lactobacillus, Leuconostoc and Bacillus. from LAB, has found practical application and is cur-

The inhibitory substance produced by Ped. pentosaceus rently being used worldwide. Nisin-producing starter is known as pediocin A and that produced by Ped. cultures have been successfully used to prevent gas acidilactici is called pediocin AcH. The molecular mass production by Clostridia in cheeses. Nisin and pediocin of pediocin AcH is 2,700 Da. The pediocin AcH mol- have a proven status as preservatives in food products. ecule has 44 amino acids. The molecule has two Pediocin is applied for protection of soft cheeses against disulphide bonds. The bacteriocin pediocin PA-1 from contamination with List. monocytogenes. ANTIMICROBIAL SUBSTANCES INCLUDING BACTERIOCINS PRODUCED BY LACTIC ACID BACTERIA 223

Nisin-producing starters were used in food applica- substances on a commercial scale to assist in food pres- tion as early as 1950s and were effective in controlling ervation. clostridial spoilage and butyric spoilage in cheeses. Nisin at the rate of 100 IU nisin/ml is also reported to REFERENCES kill lactobacilli or pediococci in brewing wort; the lac- tobacilli or pediococci are tolerant to the antimicrobial (1) Dave RI, Shah NP. 1997. Characteristics of bacteriocin pro- duced by Lactobacillus acidophilus LA-1. Int Dairy J 7: action of hops. Use of bacteriocinogenic LAB as starter 707-715. cultures to achieve pure culture vegetable fermentations (2) Hoover DG, Steenson LR (eds). 1993. Bacteriocins of Lac- has been suggested. Nisin-producing Lactococcus lactis tic Acid Bacteria, Academic Press, New York. subsp. lactis has been used to inhibit L.plantarum when (3) IDF. 1998. The use of nisin in cheese making. International Dairy Federation, Brussels, Belgium, Bulletin No. 329. grown together. Nisin producing strain of L. lactis subsp. lactis does not affect the growth of Leuconostoc (4) Klaenhammer T. 1993. Genetics of bacteriocins produced by lactic acid bacteria. FEMS Microbiol Rev 12: 39. mesenteroidres. (5) Salminen S, von Wright A (eds). 1995. Lactic Acid Bacte- Some other bacteriocins that have received indus- ria, Marcel Dekker, New York. trial attention are pediocin for control of Listeria, (6) Shah NP, Dave RI. 1999. Characteristics of bacteriocin like sakacins for specific preservation of fermented sau- inhibitory substances produced by Lactobacillus acidophi- sages, carnocins for meat and fish preservation and lus (BDLA-1, 2409 and MOLA-2), Lactobacillus lactococcins for induced lysis of starter cultures so as fermentum (5174) and Lactobacillus plantarum (2903). to control the extent of fermentation in these products Bioscience Microflora 18: 109-117. by selected LAB. (7) Tagg JR, Dajani AS, Wannamaker LW. 1976. Bacterio- cins of Gram-positive bacteria. Bacteriol Rev 40: 722-756. The recent trend is to limit or eliminate the use of (8) Vuyst L, Vandamme EJ (eds). 1994. Bacteriocins of Lac- chemical preservatives by replacing them withbio-pre- tic Acid Bacteria: Microbiology, Genetics and Application, servatives, which are believed to be less harmful than 1st ed, Blackie Academic and Professional, London and chemical preservatives. This could also pave the way New York. for future production and application of antibacterial