Bacteriocin of LAB Importance for Dairy Industry

7/28/2019 Bacteriocin of LAB Importance for Dairy Industry

1/14

Eng. Life Sci. 2012, 12, No. 4, 419432 419

Dora Beshkova

Ginka Frengova

Laboratory of Applied

Biotechnologies, Department of

Applied Microbiology, Bulgarian

Academy of Sciences, TheStephan Angeloff Institute of

Microbiology, Plovdiv, Bulgaria

Review

Bacteriocins from lactic acid bacteria:Microorganisms of potential biotechnological

importance for the dairy industryBacteriocins are a heterogeneous group of ribosomally synthesized, extracellularlyreleased,bioactive peptides or proteinsdisplayingantimicrobialactivity against otherbacteria. Over the last two decades, there has been an explosion of basic and appliedresearch on lactic acid bacteria (LAB) bacteriocins, primarily due to their potentialapplication as biopreservatives in food and food products to inhibit the growth offood-borne bacterial pathogens. Although bacteriocins can be produced in the foodmatrix during food fermentation (in situ), bacteriocins by LAB can be producedin much higher amounts during in vitro fermentations under optimal physical andchemical conditions. Because of the complexity of the food matrix and the difficultyof quantifying bacteriocin activities in foods, in vitro studies can be performed tosimulate and study the in situ functionality of bacteriocinogenic starters. In situbacteriocin production is most promising for a fast, widespread, and legal use ofbacteriocins to achieve the desirable fermentation and a safe final product. Thebacteriocin production may be of utmost importance when bacteriocin-producingLAB are added to foods as starters or protective cultures (adjunct culture). In thecurrent review, our interest is mainly focused on the research of in situ bacteriocinproduction through finding the potential of the bacteriocinogenic cultures, whichhave biotechnological importance for the dairy industry.

Keywords: Bacteriocins / Dairy industry / Fermented food / In situ production / Lactic acidbacteria

Received: December 12, 2011; revised: February 16, 2012; accepted: April 5, 2012DOI: 10.1002/elsc.201100127

1 Introduction

At present, scientificliterature and the researchcommunity have

generally adopted Klaenhammers [1] definition of bacteriocins,

i.e. bacteriocins are a heterogeneous group of ribosomally syn-

thesized, extracellularly released bioactive peptides or proteins

displaying antimicrobial activity against other bacteria.Histori-

cally, in 1976, Tagg et al. [2]defined bacteriocinsas proteinaceous

compounds, synthesized by both Gram (+) and Gram () bac-

teria, and exhibiting inhibitory activity against species closely

related to the bacteriocin producer. Information on bacteriocins

was first published in 1925, when researchers found out that a

biologically active substance produced by strain Escherichia coli

V appeared to have antagonisticactivity against another strain of

the same species (E. coliF) [3]. Later, similar antimicrobial sub-

stances produced byE. coliwere found and called colicins [4].

Correspondence: Dr. Dora Beshkova ([email protected]), In-

stitute of Microbiology, Laboratory of Applied Biotechnologies, 139

Ruski Blvd, 4000 Plovdiv, Bulgaria

Abbreviations: CFU, colony forming units; LAB, lactic acid bacteria

Bacteriocins can be produced by different species of Gram (+)

or Gram () bacteria: bacteriocins, bacillocin Bb, and pyocin Pa

produced by the soil-associated bacteria Bacillus brevis Bb and

Pseudomonas aeruginosaPa, respectively [5]; bacteriocins, aure-

ocin A 53 and aureocin A 70bystrains ofStaphylococcus aureus

[6, 7]; ruminal bacteriocinsby ruminal bacterium Streptococ-

cus bovis [8]; bacteriocinby purple nonsulfur phototrophic

bacteria, Rhodobacter capsulatus ATCC 17016 [9]; differently

called bacteriocinsby representatives of the lactic acid bacteria

(LAB) [1017]. Bacteriocin synthesis by LAB was first reportedin 1928 [18]. This biologically active substance was later chem-

ically defined as a polypeptide [19] and given the name nisin

[20, 21]. There is a growing interest in bacteriocins produced by

representatives of different LAB genera and new data have been

continuously reported.[1017] LABare widelyused in theman-

ufacturing of fermentedfoodand areamong thebest studied mi-

croorganisms. Detailed knowledge of a number of physiological

traits has opened new potential applications for these organ-

isms in the food industry, while other traits might be beneficial

for human health [22]. Owing to their typical association with

food fermentation and also their long tradition as food-grade

C 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.els-journal.com


2/14

420 D. Beshkova and G. Frengova Eng. Life Sci. 2012, 12, No. 4, 419432

bacteria, LAB are generally recognized as safe (GRAS) [23].

LAB can exert a biopreservative or inhibitory effect against

other microorganisms as a result of competition for nutrients

and/or of the production of antagonistic compounds such as

organic acids (mainly lactic acid), ethanol, aroma compounds,

fatty acids, hydrogen peroxide, bacteriocins, and nonbacteri-

ocin antibacterial compounds [24]. The persistent interest of

researchers in LAB bacteriocins is prompted by their potentialapplication as food biopreservatives, i.e. they offer a successful

prospective alternative strategy for inhibitingthe growth of food-

borne bacterial pathogens. This opens perspectives for the use

of bacteriocin-producing LAB (starters or protective cultures) in

fermented foods [2547], or of (purified) bacteriocin prepara-

tions in both fermented and nonfermented foods [27,4858] to

improve food quality, naturalness, and safety.

The fact that bacteriocins are active against numerous

foodborne and human pathogens, are produced by GRAS

microorganismsLAB, and are readily degraded by proteolytic

host systems makes them attractive candidates for biotechno-

logical applications. This review focuses on the researchs of the

production and application potential of LAB bacteriocins for thedevelopment in the biotechnology of LAB.

2 Classification and properties of LABbacteriocins

The bacteriocin family includes a wide variety of peptides and

proteins in terms of their size, microbial targets, and mecha-

nisms of action and immunity. Several attempts have been made

to classify LAB bacteriocins. Based on structural, physicochem-

ical, and molecular properties, bacteriocins from LAB can be

subdivided into three major classes [1, 59]. Nonetheless, this

classification is continuously being reviewed and it is evolvingwith the accumulation of knowledge and the appearance of new

bacteriocins [10,13,6068].The currentlyaccepted classification

by the research community is as follows:

Class I: Lantibiotics ([small], cationic, hydrophobic peptides

[


3/14


4/14


5/14

Eng. Life Sci. 2012, 12, No. 4, 419432 Bacteriocins from lactic acid bacteria 423

ing cheese manufacture and ripening. The organisms in mixed-

strain starters used in the manufacture of the cheeses belong

mainly to the genera Lactococcus(Lc. lactissubsp. lactis[Lc. lac-

tis], Lc. lactis subsp. cremoris [Lc. cremoris])mesophilic, the

best known or Lactobacillus(Lb. helveticus, Lb. delbrueckiisubsp.

bulgaricus), and Streptococcus thermophilusthermophilic, the

best known, depending on the specific application [93]. The

lactic fermentation of milk is required for cheese production.While some cheeses arestill made from nonpasteurised milk and

may even depend on the adventitious natural lactic flora (non-

starter LABNSLAB) for the fermentation, most are produced

on a commercial scale using the appropriate starter culture. The

contribution of NSLAB to the formation of the organoleptic

characteristics defining the quality of cheese is unclear. Their

presence can have both a positive and, more frequently, negative

effect, manifested in concomitant defectscalcium lactate crys-

tal formation, off-flavor development, and slit formation. One

of the most promising strategies to control NSLAB populations

is to employ well-characterized LAB, as adjunct cultures that

suppress the emergence of wild nonstarter cultures [94]. With

regards to this, the live bacteriocin-producing cultures can be asignificant alternative for improving the food safety and quality

of dairy products. The effect of in situ bacteriocin production

achieved using bacteriocinogenic dairy cultures as starter or ad-

junct cultures to obtain various types of cheese, can be expressed

by defining the role of these cultures in the following two ar-

eas: bacteriocin-producing LAB to control adventitious micro-

bial populations and as protective cultures to inhibit the growth

of pathogenic microorganisms in cheese; bacteriocin-producing

LAB as cell lysis-inducing agents to improve cheese quality and

flavordiscussed in separate sections of this article.

4 Bacteriocin-producing LAB to controladventitious microbial populations and asprotective cultures to inhibit the growthof pathogenic microorganisms in cheese

Thelacticin481-producing andlacticin3147-producing cultures

have been used successfully to improve the quality of Cheddar

cheese through the inhibition of NSLAB [28, 43]. O `Sullivan

et al. [43] observed a reduction of 4 log units in the number

of NSLAB after 4 months of ripening in experimental Cheddar,

with lacticin 481-producing strain Lc. lactisCNRZ 481 used as

an adjunct to the lactococcal starter culture Lc. lactisHP, com-

pared with the same number of bacteria in the control cheese

(obtained with the standard starter culture only). At the endof the ripening period (after 6 months), the recorded decrease

in the number of NSLAB was 2 log units. Other authors [28]

did not find NSLAB throughout the ripening period (6 months)

in experimental Cheddar cheese prepared with a mixed starter

culture comprising three strains (Lc. lactis DPC 3147, Lc. lac-

tisDPC 3204, Lc. lactisDPC 3256), isolated from natural yogurt

and producing bacteriocin, lacticin 3147. In comparison, in con-

trol, cheese produced with the commercial cheesemaking strain

Lc. cremoris DPC 4268, the number of living NSLAB cells was

107.5 CFU g1 after 4 months. The amount of bacteriocin de-

tected in cheese made with bacteriocin-producing starter was

approximately 1280 AU mL1, which remained stable over the

entire ripening period.

Insitu production of nisinZ byLc. diacetylactisUL 719,cocul-

tivated with Lc. cremorisKB and Lc. lactis(at a ratio of 1.0:1.5:1.5

vol.) in Cheddar cheese, was determined throughout ripening

(6 months) [39]. Nisin concentration of 306 IU mL1 was ob-

tained. In addition to the nisin-producing activity of the mixed

starter, researchers evaluated the antagonistic effect of bacteri-ocin against the growth of Listeria innocua. A reduction of 3.0

log units in the level of living cells of the pathogen was found

in experimental cheese produced with encapsulated nisin, and a

reductionof 1.5log units in experimental cheese obtained with a

nisinogenic starter. At the end of ripening process in cheese pro-

duced with a nisin preparation, about 10 CFU g1 ofL. innocua

and90% of theinitial activity of nisin wasestablished, compared

to 104 CFU g1 and 12% for cheese produced with a nisinogenic

starter. The use ofLc. diacetylactisUL 719 in a mixed starter cul-

ture did not appear to be the best strategy to inhibit L. innocua

in Cheddar cheese. However, a mixed culture containing a nisin

Z-producing strain might be used for controlling postcontami-

nant organisms, as they are usually present in low numbers. Thedata obtained by the authors on nisin activity in experimental

Cheddar cheese produced witha nisinogenic starter (Lc. diacety-

lactisUL 719) aresimilar to those reported previously forGouda

cheese [35]. Bauksaim et al. [35] reported that Gouda cheese of

good qualityandsafe toeat canalsobe obtained bythe aforemen-

tioned nisin-producing culture Lc. diacetylactisUL 719, when it

is addedto commercialFloraDanicastarterin a ratioof 0.6:1.4%.

In the experimental cheese, a maximum nisin concentration of

512 IU g1 was recorded (after 6 weeks), followed by a decrease

in the activity to 128 IUg1 after 27 weeks, and up to 32 IU g1

after 45 weeks of ripening. Maisnier-Patin et al. [25] evaluated

the potential of another nizin-producing starter Lc. lactisCNRZ

150 to inhibit Listeria monocytogenesduring Camembert cheese

manufacture and ripening. Maximum nisin concentration of ca700IUg1 was obtained in curd at 9.0h (during the exponential

phase of growth of bacteriocin-producing culture), then nisin

concentrations decreased slowly during 924 h, and dramati-

cally to ca 200 IU g1 during ripening. In the presence of nisin,

the numbersofL. monocytogenesdecreased rapidlyfrom6 to24 h

and a difference of 2.4 log CFU g1 between numbers of Liste-

ria in cheeses made with Nis+ (Lc. lactisCNRZ 150) and Nis

(Lc. lactisCNRZ 1076) starter cultures was maintained through-

out ripening (6 weeks). Another nisin Z-producing strain Lc.

lactisIPLA 729 has been successfully applied on the inhibition

of the spoilage strain ofClostridium tyrobutiricum CECT 4011, a

lateblowingagent,in semi-hard Vidiagocheesemaking[44]. The

authors have obtained and studied three cheeses: experimental

cheesenisinogenic culture was co-cultivatedwith a mesophilic

starter IPLA-001 composed of Lc. diacetylactisIPLA 8381 and

Leuconostoc citreum IPLA 616; control cheeseinstead of the

nisin Z-producing culture, an acidifying nonbacteriocinogenic

strain Lc. lactis IPLA 947 was used in the mixed starter; com-

mercial cheesethe nisin producer was replaced by KNO 3 as

a gas-blowing preventing agent. A maximum concentration of

1600 IU nisin mL1 was measured in the experimental cheese

betweenday 1 andday15 of theripeningprocess, whileat theend

of the process (30 days), the activity decreased. During ripeningof thethreetypes of cheese, thefollowing trendsin thegrowth of



6/14


clostridiawere observed, and the following values for the number

of live cells were determined: the number decreased from 1.2

106 to 1.3 103; the number increased to 3.5 107; the num-

ber increased to 1.99 109 for the experimental, control, and

commercial cheese, respectively. The authors concluded that the

inclusion of the nisin-producing strain IPLA 729 in the mixed

starter IPLA-001 made it possible to prepare a balanced cul-

ture that will be used in the making of Vidiago cheese, therebyimproving the ability to suppress spoilage microorganisms and

contributing to better organoleptic properties in comparison

with commercial starters.

Onepossibilityfor the improvementof themetabolic produc-

tivityof an microorganisms is genetic modification.This strategy

wassuccessfully used by Ryan et al.[28] as follows: a bacteriocin-

producing transconjugant Lc. lactisDPC 4275 was obtained by

transferring a 63-kb plasmid, pMRC01 (encodes bacteriocin,

lacticin 3147 production) from strain Lc. lactis DPC 3147 to

the commercial Cheddar cheesemaking starter Lc. cremorisDPC

4268; the obtained transconjugant was defined as Bac+ and

Imm+, i.e. possessing properties of lacticin 3147 production and

immunity to lacticin 3147. Furthermore, on the one hand, thetransconjugant had retained the lacticin 3147-producing activ-

ity of the parent strain Lc. lactisDPC 3147, while, on the other

hand, it maintained the original characteristics of the commer-

cial starter strain, i.e. theproduced lactic acid levels were similar.

Throughout ripening (6 months), the concentration of bacteri-

ocin, lacticin 3147, determined in Cheddar cheese (made with a

transconjugant strain as a single-strain starter) did not change

and this correlated with a significant decrease in the number

of NSLAB. Later, the lacticin 3147-producing transconjugant

Lc. lactisDPC 4275 was used by other authors as a component

of a starter culture not only for obtaining reduced fat cheddar

[34], but also for obtaining Cottage cheese [33]. At both ripen-

ing temperatures (7Cand12C), theNSLAB populations in the

corresponding cheeses made with the lacticin 3147-producingculture, Lc. lactisDPC 4275, increased much more slowlyduring

ripening andreachedmarkedly lower numbers (ca103 CFUg1)

at theend (240 days)of theripening[34]). Inagreement with the

observations of Folkertsma et al.[95] forfull fatCheddar cheese,

elevation of ripening temperature from 7C to 12C resulted in

a more rapid developmed of NSLAB (to ca 107 CFU g1) in

cheeses made with the nonbacteriocinogenic mixed starter ([Lc.

lactisDPC 4268+ Lc. cremorisDPC4269] or Lc. lactisDPC 4268

alone). The authors concluded that the use of the bacteriocino-

genic Lc. lactisDPC 4275 can reduce the risk of flavor defects in

reduced fat cheese, especially when ripening is at high tempera-

ture, and shouldenable theproductionof cheeses with morepre-

dictable and consistent flavor. The effectiveness of lacticin 3147

as a naturalpreservative wasdetermined in Cottage cheese that

was subsequently inoculated with approximately 104 L. mono-

cytogenesScott A g1 [33]. During the first week of storage (at

4C), in the experimental cheese (made with the bacteriocin-

producing transconjugant Lc. lactisDPC 4275 as a starter), the

concentration of lacticin 3147 was determined to be 2560 AU

mL1. Furthermore, a decrease of 99.9% was registered in the

number of L. monocytogenes in the same cheese within 5 days,

whereas in the control cheese (made with commercial starter Lc.

lactisDPC4268), theinitial numberof inoculated pathogencells

remained essentially unchanged. Another genetically modified

starter culture Lc. lactisMM 217, capable of producing pediocin

in situ, was used as a single starter culture to improve the mi-

crobiological safety of Cheddar cheese precontaminated with

L. monocytogenes[32]. The nonbacteriocinogenic starter culture

Lc. lactisMM 210was used as an recipient forplasmid (pMC117)

coding for pediocin PA-1 production. The electrotransformed

derivate of strain MM 210 containing pMC117, was named Lc.

lactis MM 217. This strain retained the growth characteristicsand acid-producing activity of the parent strain. At the end of

theripeningprocess of cheeses (6 months at 8C),the number of

live L. monocytogenescellsdecreasedfrom3.5logCFUg1 (initial

concentration) to about 1.0 log CFU g1 in experimental cheese

(made with a bacteriocin-producing starter culture), compared

to a recorded number of pathogen cells of 4.0 log CFU g1

in the control cheese (made with a nonbacteriocin-producing

starter culture). The level of pediocin activity decreased from

approximately 64,000 AU g1 after 1 day to 2000 AU g1 after 6

months. Rodriguez et al. 2005 [46] evaluated the antimicrobial

activity of bacteriocin-producing transformants (Lc. lactis CL

1 [Ped+]-pediocin-producing strain and Lc. lactis CL 2 [Nis+,

Ped

+

] nisin-and pediocin-producing strain) against L. monocy-togenes, S. aureus, and E. coliO157:H7 during cheese ripening.

The wild strains Lc. lactisESI 153 and Lc. lactisESI 515(Nis+)

were selected by their technological and/or antimicrobial prop-

erties, used as starter cultures in cheese manufacture and were

transformed to produce pediocin PA-1, as reported earlier [96].

Pediococcus acidilactici347 (Ped+), Lc. lactis ESI 153, Lc. lactis

ESI 515 (Nis+), and their respective pediocin-producing trans-

formants Lc. lactisCL 1 (Ped+) and Lc. lactisCL 2 (Nis+, Ped+)

were added individually as adjunct to the commercial starter

culture MA 016. In the experimental cheeses containing one of

the above bacteriocinogenic cultures, bacteriocin activity was

established during their ripening (30 days). After 30 days, in the

presence of the bacteriocinogenic cultures Lc. lactisCL 1 (Ped+)

or Lc. lactisCL 2 (Nis+, Ped+), there was a certain decrease inthe L. monocytogenescounts by 2.97 and 1.64 log units, S. aureus

by 0.98 and 0.40 log units, and E. coliO157:H7 by 0.84 and 1.69

log units when compared with control cheese (made without

adjunct culture). The combination of the bacteriocins nisin and

pediocin, synthesized by Lc. lactisCL 2 (Nis+, Ped+) in the re-

spective experimental cheeses, revealed a higher inhibitory effect

only against the pathogen E. coli. All cheeses investigated by the

authors were inoculated with each of the given pathogen in an

approximateconcentrationof 6 log CFU mL1,whichwashigher

than the possible contamination of natural milk. Similarly Zot-

tola et al.[27] suggestedthat theuse of bacteriocinogenic culture

as an ingredientin pasteurized process cheese or coldpack cheese

spreads couldbe an effectivemethod of controllingthe growthof

undesirable microorganisms in these processed foods. A nisin-

producing transconjugant Lc. cremoris JS 102 was added as an

adjunct to lactoccocal starter Lc. lactisNCDO 1404 in Cheddar

cheesemaking, which contained between 400 and 1200 IU of

nisin g1 cheese. The shelf life (at 22C) of the nisin-containing

cheese was significantly greater than that of the control cheese

(the cheeses were inoculated with 2000 spores of Cl. sporogenes

PA 3679 during manufacture). Significant reduction in numbers

of the nonspore-forming test microbes (L. monocytogenes V 7

and S. aureus 196 E) were observed in experimental cheeses at

both temperatures (22C and 37C).



7/14


Figure 1. Overview of potential application ofLAB-producing bacteriocins.

Besides lactococci as adjunct cultures during in situ bacteri-

ocin production, the quality of the cheese may be improved by

also using lactobacilli [36,47] and enterococci [26,29,40]. Ryan

et al. [36] applied one strategy for manipulation of cheese flora

using combinations of lacticin 3147-producing and -resistant

cultures. A stable, more resistant Lb. paracaseiDPC 5337 (which

was 32 times less sensitive to bacteriocin, lacticin 3147 than

parental strain Lb. paracaseiDPC 5336) was used as adjunct cul-

tures in two separatetrialsusingeither Lc. lactisDPC3147(anat-

ural producer) or Lc. lactisDPC 4275 (a lacticin 3147-producing

transconjugant) as the starter. These lacticin 3147-producing

starters were previously shown to control adventitious NSLABin cheddar cheese [28].Lacticin-3147activity wasassayed during

manufacture and reached approximately 12802560 AU g1 of

cheese. This level of activity was maintained throughout ripen-

ing and correlated with a reduction in the growth rate of NSLAB

in the control cheeses. The authors deducted that the resistant

adjunct strain formed the dominant Lactobacillus population

(levels of 107 CFU g1, in contrast to the sensitive strains

levels 100- to 1000-fold lower) in the experimental cheeses that

were with improved quality and with the bitter flavor being al-

most undetectable. Bogovic Matijasic et al. [47] implemented

a model for inhibiting the growth of Cl. tyrobutyricum, with

the pathogen inoculated in advance (at a concentration of 2.5

103

spores mL1

) in semi-hard cheese, obtained by using abacteriocinogenic culture (Lb. gasseriK 7) as an adjunct to the

commercial starter S. thermophilusTH4DVS. In the experimen-

tal cheese, the bacteriocin-producing culture did not inhibit the

thermophilic starter, but reduced the number of the nonstarter

mesophilic lactobacilli to about 100CFU g1 throughout the en-

tire ripening (8 weeks). This inhibitory effect was observed de-

spite thefact that nobacteriocins were found in theexperimental

cheese. In addition, the pH values and concentrations of or-

ganic acids (factors contributing to the antimicrobial properties

of LAB) were similar in the cheeses produced in the presence

or absence of a bacteriocin-producing culture. The authors sug-

gest the following possible explanations of this phenomenon:

nonuniform bacteriocin distribution in cheese, adsorption to

the caseins in the curd, or bacteriocin degradation by intracel-

lular proteases, as reported earlier by other authors [97]. Villani

et al. [26] used Enterococcus faecalis226 NWC (culture produced

the bacteriocin, enterocin 226 NWC) as a starter in Mozzarella

cheesemaking from water buffalo milk. When L. monocytogenes

was cocultured with a bacteriocin-producing culture in recon-

stituted skim milk, the live cells of the pathogen decreased to a

level of 1.5 107 and were completely destroyed after 7 and 72 h

of incubation of the mixed culture, respectively. For compari-son, the number of live cells of the pathogen was 9 108 in

the development of L. monocytogenesas a monoculture. Later,

Nunez et al. [29] reported that another enterocin-producing

strain Ent. faecalis INIA 4 was successfully used for Manchego

cheesemaking from raw ewes milk. Listeria monocytogenesOhio

counts decreased by 3 log units after 8 h, and by 6 log units

after 7 days in cheese made with an enterocinogenic culture,

whereas no inhibition was recorded after 60 days in control

cheese made with commercial starter (Lc. cremoris+ Lc. lactis).

Anotherstrain, L. monocytogenesScottA, wasnotinhibited in the

presence of a bacteriocin-producing culture, used either alone

or as an adjunct to a commercial starter during cheese manu-

facture, but decreased by 1 log unit and 2 log units, respectively,after 7 and 60 days of ripening. The values of enterocin activi-

ties, determined 8 h after making theexperimental cheeses (with

a mixed starterbacteriocinogenic + commercial strains, and

with a single starterbacteriocinogenic culture), were 2000

3000 AU g1 and 40006000 AU g1, respectively. These re-

sults are in agreement with bacteriocin production by Ent. fae-

cium 7C5 under Taleggio cheesemaking conditions, which still

occurred at 25 h andpH 4.9[98]. Sarantinopouloset al.[40]have

investigated the possibility of the use of bacteriocinogenic strain

Ent. faecium FAIR-E 198 as a adjunct culture to the traditionally



8/14


used mixed starter (S. thermophilusACA-DC 7, Lc. lactisACA-

DC 52, and Lb. bulgaricus ACA-DC 84) to manufacture Greek

Feta cheese. This strain was isolated from Greek Feta cheese and

demonstrated an antagonistic effect against Listeria.

5 Bacteriocin-producing LAB as celllysis-inducing agents to improve cheesequality and flavor

Another important aspect of using bacteriocinogenic cultures

is the possibility to induce controlled lysis of a LAB starter cul-

ture or NSLAB in their presence, and subsequent intracellular

release of proteinases and peptidases, resulting in rapid onset

of proteolysis, i.e. a new alternative is proposed for acceleration

of cheese ripening aimed at obtaining dairy products with im-

proved organoleptic characteristics [30,31,37,38,4143,45,99].

Morgan et al. [30] described a method for increasing the rate

of lysis of the commercial starter culture Lc. cremorisHP during

ripening of Cheddar cheese by adding the bacteriocinogenic cul-ture Lc. lactisDPC3286 (encodesthe synthesis of lactococcins A,

B, M)to thestandardlactococcal starter.In a laboratory-scale sys-

tem, it was found that the LHD (lactate dehydrogenase) activity,

determined onday180 (end ofthe ripeningprocess) inthe exper-

imental cheese (made with bacteriocinogenic adjunct culture),

was 66% higher than the activity in the control cheese (obtained

with the lactococcal starter). In addition, higher values were also

recorded for both enzymesglucose-6-phosphate dehydroge-

nase and postproline dipeptidyl aminopeptidase in the experi-

mental cheese, resulting in higher concentrations of free amino

acids by 47% than those in the control cheese. With the suc-

cessful application of the mixed culture (Lc. cremorisHP+ bac-

teriocinogenic adjunct strain Lc. lactisDPC 3286), the authors

obtained a dairy product of improved quality, reduced bitter-ness, and higher grading scores. The use of the two-component

culture on a pilot scale, however, created a problem by killing

the acid-producing strain included in the mixed starter in the

making of Cheddar cheese. Later, the same authors (41) resolved

this problem by applying a new three-component mixed culture

consisting of the following cultures: a lactococcin A, B, and M

strain producer (Lc. lactisDPC 3286), possessing the ability to

lyse thesecondculture(Lc.cremorisHPsensitiveto lactococcin

A, B, and M activity), and a third culture ( S. thermophilusDPC

1842acid producing and resistant to lactococcin A, B, and M

activity) for pilot-scale Cheddar cheesemaking. In the experi-

mental cheese (made with a bacteriocinogenic adjunct culture),

higher (approximately two times) concentrations of free aminoacids were recorded, higher release rates of intracellular LDH

(by 265%), and a decrease in bitterness compared to the con-

trol cheese (made with Lc. cremoris HP alone or with a mixed

starterLc. cremorisHP+ S. thermophilusDPC 1842). Another

bacteriocinogenic strain, Lc. lactis CNRZ 481, producing lac-

ticin 481, was also used as an adjunct to the commercial starterculture Lc. lactisHP for pilot-scale Cheddar cheesemaking, but

without impeding the production of the amount of acidrequired

for this type of cheese [43], in contrast to lactococcin A, B, and

M strain-producer Lc. lactis DPC 3286, which was reported to

have a negative effect in this context [41]. In the experimental

cheese (made with a mixed starterbacteriocinogenic culture

+ commercial starter culture), 45 times higher levels of LDH

were determined compared with those in the control cheese

(made with a commercial starter culture alone). Other authors

[45] also reported that addition of a bacteriocinogenic strain

Lc. lactisINIA 415 (strain containing the structural gene encod-

ing lacticin 481 and nisin Z production) as adjunct culture to

commercial S. thermophilus TA 052 and Lc. lactis INIA 415-2(a nonbacterioconogenic mutant) is a successful alternative for

the acceleration of proteolysis of Hispanico cheese. Streptococcus

thermophilusTA 052 counts were lower (about 1 log unit) in ex-

perimental cheese (made with bacteriocinogenic culture) on day

15 of ripening. From day 25 to day 75 (end of ripening), in the

presence of the bacteriocinogenic culture, i.e. in the experimen-

tal cheese, the total free amino acid concentrations were about

2.5 times higher than those in the control cheese (made without

a bacteriocinogenic adjunct culture). Later, Garde et al. [99] de-

scribed another alternative for Hispanico cheesemaking using a

lactococcal mixed starter (lacticin 481-producing Lc. lactisINIA

639 + lacticin 481-nonproducing Lc. lactisINIA 437) to which

a lactobacillus strain (Lb. helveticus LH 92) was added, whichis sensitive to lacticin 481 and possesses high aminopeptidase

activity. In the control cheese, about a twofold lower rate of pro-

teolysis was recorded, and about 1.8 times lower values for the

activity of cell-free aminopeptidase was determined compared

with the values obtained in the experimental cheese after 25

and 7 days of ripening, respectively. After 25 days, in the experi-

mental cheese, the concentrations of total free amino acids were

determined to be about 2.3 times higher than those in the con-

trol cheese. As a result of using the lacticin 481-producing cul-

ture, the cheese obtain by the authors had improved quality and

reduced bitterness. Martinez-Cuesta et al. [37] studied the po-

tential of lacticin 3147-producing transconjugant Lc. lactisIFPL

3593 (used as a starter) toinduce lysis of thetwo cultures (Lc. lac-

tisT1and Lb.caseiIFPL 731bothshowedhigh aminopeptidaseactivity)addedto thestarter, thusachieving accelerated processes

of proteolysis and ripening in semi-hard cheese, accompanied

by a parallel significant increase in its sensory characteristics

intensity of aroma and cheese taste. The bacteriocin-producing

transconjugant Lc. lactisIFPL 3593 was obtained by transferring

a 46-kb plasmid, pBAC 105 (encodes bacteriocin, lacticin 3147

production) from strain Lc. lactis IFPL 105 to the commercial

cheesemaking starter Lc. lactisIFPL 359. The transconjugant was

defined as Bac+ and Imm+, i.e. possessing properties of lacticin

3145 production and immunity to lacticin 3147. The values of

X-prolyl-dipeptidyl aminopeptidase activity were significantly

higher (about two times) in experimental cheese (made with the

bacteriocinogenic culture, as starter). In addition, detection of

intracellular activity and loss of cellular viability of starter ad-

juncts (Lc. lactisT1 and Lb. caseiIFPL 731) were simultaneous.

The concentration of amine nitrogen in experimental cheese on

day 45 (end of ripening) was 16% higher than in the control

cheese (made with nonbacteriocinogenic parental strain Lc. lac-

tisIFPL 359 [commercial starter] and adjuncts cultures [Lc. lactis

T 1 + Lb. caseiIFPL 731).

Enterococcus species also were added as a bacteriocin-

producing adjunct to commercial starter culture in cheese

making [31, 38]. A feasible and a cost-effective method for

increasing the rate of starter lysis during semi-hard Hispanico



9/14


Table 2. Bacteriocin-producing LABdirection for their use in different cheesemaking.

Bacteriocin-producing culture Bacteriocin Directions for application Observed effects Reference

1 2 3 4 5

Lc. lactisDPC 3147 Lc. lactisDPC

3204 Lc. lactisDPC 3256

Lacticin 3147 As mixed starter culture in Cheddar

cheesemaking

No NSLAB detected in experimental

cheese at the end ripening 6

months

[28]

Transconjugant Lc. lactisDPC 4275 Lacticin 3147 As single-strain starter culture inCheddar cheesemaking

A significant reduction in the levelsof NSLAB

[28]

Transconjugant Lc. lactisDPC 4275 Lacticin 3147 As single-strain in reduce fat

Cheddar cheesemaking

A reduction in the number of NSLAB

to 103 CFU g1 (at both ripening

temperature 7C and 12 C at the

end ripening (240 days) in

experimental cheese

[34]

Transconjugant Lc. lactisDPC 4275 Lacticin 3147 As single-strain starter in Cottage

cheese

A 99.9% reduction in the counts of L.

monocytogeneswithin 5 days at

4C in experimental cheese

[33]

Lc. lactisDPC 3147 (natural

producer) Lc. lactisDPC

(transconjugant)

Lacticin 3147 As starter culture added individually

to Lb. paracaseiDPC 5337,

resistant to lacticin 3147

A manipulation of cheese flora [36]

Transconjugant Lc. lactis3593 Lacticin 3147 As starter culture to starter adjuncts

(Lc. lactisT1 and Lb. caseiIFPL 731

in semi-hard cheesemaking

An increase in values of amino

peptidase activity (about two

times) and amine nitrogen (16%higher) at the end ripening (42

days)

[37]

Lc. lactisCNRZ 481 Lacticin 481 As adjunct to the lactococcal starter

Lc. lactisHP in Cheddar

cheesemaking

A 2 log units reduction n the counts

of NSLAB in experimental cheese

at the end of ripening (6 months);

an increase in LDH levels (fivefold

higher); improve the quality of

cheese

[43]

Lc. lactisINIA 639 Lacticin 481 As starter along with Lc. lactisINIA

437 and Lb. helveticusLH 92 in

Hispanico cheesemaking

An increase in proteolysis (to twofold

higher); an increase in values of

amino peptidase activity (to

1.8-fold higher) and total amino

acids (2.3-fold higher) after 25

days; a reduction in bitterness.

[99]

Lc. lactisINIA 415 containing the

gene encoding lacticin 481 and

nisin production

Lacticin 481+ nisin Z As adjunct to the commercial culture

S. thermophilusTA 052 and Lc.

lactisINIA 415-2 (a

nonbacteriocinogenic mutant) in


An increase in secondary proteolysis

and levels of total free amino acid

(1.49 and 2.34-fold higher,

respectively) on day 75.

[45]

Lc. lactisDPC 3286 Lactococcin A, B, M As adjunct to the lactococcal starter

Lc. cremorisHP in Cheddar

cheesemaking

An increase in proteolysis; An

increase in concentrations of total

free amino acids; A reduction in

bitterness and a cheese with

improved flavor and quality.

[30]

Lc. lactisDPC 3286 Lactococcin A, B, M As adjunct to the starter Lc. cremoris

HP + S. thermophilusDPC 1842 in

Cheddar cheesemaking

An increase in LDH levels of 265%;

An increase in level of total free

amino acids (about two times); a

reduction in bitterness.

[41]

Lc. diacetylactisUL 719 Nisin Z As starter coculture in Cheddar

cheesemaking

A reduction in the counts ofL.

innocuato 104 CFU g1 in

experimental cheese at the end of

ripening (6 months)

[39]

Lc. lactisIPLA 729 Nizin Z AS adjunct to the mesophilic starter

IPLA 501 (Lc. diacetylactis+

Leuconostoc citreum IPLA 616) in

semi-hard Vidiago cheesemaking

A successfully control of the growth

ofCl. tyrobutiricum in

experimental cheese; A

gas-blowing preventing agent

[44]

Lc. lactisCNRZ 150 Nizin As starter culture together with Lc.

lactisCNRZ 1076 in Camembert

cheesemaking

A 2.4 log CFU g1 reduction in the

numbers ofL. monocytogenes

(throughout ripening6 weeks)

[25]

Transconjugant Lc. cremorisJS 102 Nisin As adjunct to the lactococcal starter

Lc. lactisNCDO 1404 in Cheddar

cheesemaking

A significant reduction in the counts

ofL. monocytogenesand S. aureus

during storage at 23C and 37C.

[27]



10/14


Table 2. Continued

Bacteriocin-producing culture Bacteriocin Directions for application Observed effects Reference

1 2 3 4 5

Electrotransformant Lc. lactis

MM217

Pediocin PA-1 As single-strain starter in Cheddar

cheesemaking

A reduction in the number of L.

monocytogenesto 1.0 log g1 at the

end of ripening (6 months) at 8C.

[32]

Transformants Lc. lactisCL1 (Ped+)and Lc. lactisCL2 (Nis+, Ped+)

Pediocin As adjunct (added individually) tothe commercial starter MA016 in

cheesemaking

A reduction in the counts ofpathogens as follows: L.

monocytogenesto 1.64 log Units;

S. aureusto 0.40 log Units and E.

colito 0.84 log Units on the end

of ripening (30 days).

[46]

Lb. gasseriK7 Bacteriocin As adjunct to the commercial starter

culture S. thermophilusTH4DVC

in semi-hard cheesemaking

Inhibition ofCl. tyrobutyricum; A

reduction in the counts of NSLAB

to< 100 CFU g1 throughout the

entire ripening (8 weeks)

[47]

Ent. faecuim FAIR-E 198 Enterocin As adjunct to the coculture starter (S.

thermophilusACA-DC7+ Lc.

lactisACADC 52+ Lb. bulgaricus

ACA-DC 8) in Greek Feta

cheesemaking

Inhibition ofListeria [40]

Ent. faecalis226 Enterocin 226 NWC As star ter culture in Mozzarelacheesemaking

Inhibition ofL. monocytogenes [26]

Ent. faecalisINIA 4 Enterocin 4 As adjunct to the commercial starter

(Lc. cremoris+ Lc. lactis) in

Manchego cheesemaking

Inhibition ofL. monocytogenesOhio

but not ofL. monocytogenesScott A

[29]

Ent. faecalisINIA 4 Enterocin 4 As adjunct to the commercial

mesophilic CH-N01-type starter

(Lc. lactis+ Lc. diacetylactis+ Lc.

cremoris+ Leuconostoc cremoris)

in semi-hard Hispanico

cheesemaking.

An increase in proteolysis; An

increase in levels of

aminopeptidase activity (about

eightfold higher) on day 15.

[31]

Ent. faecalisINIA 4 Enterocin 4 As adjunct to the commercial

mesophilic LD-type starter (Lc.

cremoris+ Lc. diacetyactis+

Leuconostoc cremoris) in semi-hard


An increase in proteolusis (1.8-fold

higher) and levels of total free

amino acids (2.17-fold higher); A

reduction in level of hydrophobic

peptides and bitterness in

experimental cheese at the end of

ripening (45 days)

[38]

cheese ripening, anda more rapid developmentof thecharacter-

istic cheese flavor has been reported [31]. These positive effects

were achievedby the authors by usingthe bacteriocin-producing

strain Ent. faecalisINIA 4 (at a low inoculation level0.003%

v/v) to the commercial LD-type (cultures producing diacetyl

and carbon dioxide) mesophilic starter culture CH-N01, con-

sisting of Lc. lactis, Lc. lactis biovar. diacetylactis, Lc. cremoris,

Leuconostoc mesenteroides subsp. cremoris to make Hispanico

cheese. From 3 to 15 days, released aminopeptidase generally

double in experimental cheeses (with bacteriocinogenic culture,

as adjunct), reaching values for activity on Lys-pNA and Leu-

pNA up to 9.8- and 6.4-fold higher, respectively, than in control

cheese. Similarly, Oumer et al. [38] concluded that early lysis of

starter cells in Hispanico cheese made from mixture of cows`and

ewes` milks (4:1) inoculated with 1.0 g bacteriocin-producing

culture Ent. faecalisINIA4kg1 wasfollowed by a higher produc-

tion of free amino acids and some volatile compounds (diacetil,

3-methyl-1-butanal)that are important for the organolep-

tic characteristics of cheese. The commercial starter (LD-type

mesophilic starter culture CH-N01 consisting of Lc. lactisbio-

var. diacetylactis, Lc. cremoris, L. mesenteroidessubsp. cremoris)

lost viability more rapidly in experimental cheese (made with

the bacteriocinogenic strain), which reached counts of up to 6

107 CFU g1 during ripening. At the end of ripening pe-

riod (45 days), the degree of proteolysis and concentrations of

total amino acids in experimental cheese was 1.80- and 2.17-

fold higher than the respective values in control cheese (absence

of the bacteriocinogenic strain). The aminopeptidase activity

increased significantly (twice) as a result of adding bacteriocino-

genic culture to milk. Inoculating milk with Ent. faecalis INIA

4 reduced the level of hydrophobic peptides that are associated

with bitterness in the experimental cheese.

6 Future prospects

The analysis of data from research work on in situ bacteri-

ocin production could not exclude any mention of the potential

for application of bacteriocin-synthesizing lactic acid cultures

(LAB), which are of great biotechnological importance for the



11/14


Practical application

The information summarized here can lead to the conclu-sion that there is a great variety of cost-effective ways thatcan be implemented in using bacteriocin-producing cul-tures as starters, as adjunct cultures for fermented foods

and as protective cultures to the surface of food products.Despite the large variety, they all possess potential prop-erties for improving food quality and safety, and are anantimicrobial alternative along the microbe food chain.

The knowledge about in situ bacteriocin production bylactic acid bacteria (LAB) can be used both to improve thelong-known applications of these microorganisms and tocreate new aspects of applications of these cultures in thefood industry. Some properties of LAB can be the key linkto the health effects in nutritious foods. For some of thephysiological properties found in LAB, it has been provedthat these organisms areexcellentfor progressive research.Moreover, LAB have been increasingly used as a model

organism for future physiological and genetic research.

dairy industry. Figure 1 schematically represents the possible

potential applications of LAB bacteriocins, and Table 2 shows

directions for using bacteriocinogenic cultures in making milk

products (different types of cheese) on the basis of the specific

data reported on this. The summarized information in Table 2

can lead to the conclusion that there is a great variety of cost-

effective ways that can be implemented in using bacteriocin-

producing cultures as starters, as adjunct cultures for fermented

foods, and as protective cultures to the surface of food products.

Despite the large variety, they all possess potential properties for

improving food quality and safety, and are an antimicrobial al-

ternative along the microbe foodchain. Bacteriocin-synthesizingLAB are preferred in their role of natural biopreservatives in

food. Biological preservation implies a novel scientifically based

approach to improve the microbiological safety of foods and is

todays response to the evergrowing consumer interest in nat-

ural foods without chemical preservatives. In conclusion, the

efforts of researchers should be directed toward selection of new

LAB strains whose features satisfy both the relevant technologi-

cal requirements for a standard starter culture in making dairy

products, as well as producing bacteriocins with antimicrobial

activity, acting as "bioconservatives," and providing quality and

safe food products. In this respect, the successful strategies will

include genetic engineering to transfer genes encoding a specific

bacteriocin production from nonstarter LAB strains to indus-trial strains of starter cultures, while maintaining their origi-

nal technological features to obtain a quality end product. The

knowledge about in situ bacteriocin production by LAB can be

used both to improve the long-known applications of these mi-

croorganisms and to create new aspects of applications of these

cultures in the food industry. Some properties of LAB can be the

key link to the health effects in nutritious foods. For some of the

physiological properties found in LAB, it has been proved that

these organisms are excellent for progressive research. Moreover,

LAB have been increasingly used as a model organism for future

physiological and genetic research.

The authors have declared no conflict of interest.

7 References

[1] Klaenhammer, T. R., Genetics of bacteriocins produced by

lactic acid bacteria. FEMS Microbiol. Rev. 1993, 12, 3986.[2] Tagg, J. R., Dajani, A. S., Wannamaker, L. W., Bacteriocins of

gram-positive bacteria. Bacteriol. Rev. 1976, 40, 722756.

[3] Gratia, A., Sur un remarquable example d`antagonisme entre

souches de colibacille. C. R. Soc. Biol. 1925, 93, 10401041.

[4] Cascales, E., Buchanan, S., Duche, D., Kleanthous, C. et al.,

Colicin biology. Microbiol. Mol. Biol. Rev. 2007, 71, 158229.

[5] Saleem, F., Ahmad, S., Yagoob, Z., Rasool, A. S., Comparative

study of two bacteriocins produced by representative indige-

nous soil bacteria. Pak. J. Pharm. Sci. 2009, 22, 252258.

[6] Netz, D. J. A., Pohl, R., Beck-Sickinger, A. G., Selmer, T. et al.,

Biochemical characterisation and genetic analysis of aureocin

A53, a new, atypical bacteriocin from Staphylococcus aureus.

J. Mol. Biol. 2002, 319, 745756.[7] Netz, D. J., Sahl, H. G., Marcelino, R., dos Santos Nascimento,

J. et al., Molecular characterisation of aureocin A 70, a multi-

peptide bacteriocin isolated from Staphylococcus aureus.J. Mol.

Biol. 2001, 311, 939949.

[8] Russell, J. B., Mantovani, H. C., The bacteriocins of ruminal

bacteria and their potential as an alternative to antibiotics.

J. Mol. Microbiol. Biotechnol. 2002, 4, 347355.

[9] Lee, S. S., Oh, T. J., Kim, J., Kim, J. B. et al., Bacteriocin from

purple nonsulfur phototrophic bacteria, Rhodobacter capsula-

tus. J. Bacteriol. Virol. 2009, 39, 269276.

[10] Cleveland,J., Montville,T. J.,Nes, I. F., Chikindas,M. L., Bacte-

riocins: safe, natural antimicrobials for food preservation. Int.

J. Food Microbiol. 2001, 71, 120.

[11] Chen, H., Hoover, D. G., Bacteriocins and their food applica-tions. Compr. Rev. Food Sci Food Safety2003, 2, 8298.

[12] Galvez, A., Abriouel, H., Lucas Lopez, R., Ben Omar, N.,

Bacteriocin-based strategies for food biopreservation. Int. J.

Food Microbiol. 2007, 120, 5170.

[13] Ennahar, S., Sashihara, T., Sonomoto, K., Ishizaki, A., Class IIa

bacteriocins: biosynthesis, structure and activity. FEMS Micro-

biol. Rev. 2000, 24, 85106.

[14] Sablon, E., Contreras, B., Vandamme, E., Antimicrobial pep-

tides of lactic acid bacteria: mode of action, genetics and

biosynthesis. Adv. Biochem. Eng. Biotechnol. 2000, 68, 2160.

[15] De Vuyst, L., Leroy, F., Bacteriocins from lactic acid bacteria:

production, purification and food applications. J. Mol. Micro-

biol. Biotechnol. 2007, 13, 194199.

[16] Cotter, P. D., Hill, C., Ross, R. P., Bacteriocins: Developing

innate immunity for food. Nat. Rev. Microbiol. 2005, 3, 777

788.

[17] Nes, I. F., Yoon, S. S., Diep, D. B., Ribosomally synthesized

antimicrobial peptides (bacteriocins) in lactic acid bacteria: A

review. Food Sci. Biotechnol. 2007, 16, 675690.

[18] Rogers, L. A., The inhibitory effect of Streptococcus lactis on

Lactobacillus bulgaricus. J. Bacteriol. 1928, 16, 321325.

[19] Whitehead, H. R., A substance inhibiting bacterial growth,

produced by certain strains of lactic streptococci. Biochem J.

1933, 27, 17931800.



12/14


[20] Mattick, A. T. R., Hirsch, A., Furter observations on an in-

hibitory substance(nisin)from lacticstreptococci. Lancet1947,

2, 57.

[21] Hurst, A., Nisin, in: Perlman, D., Laskin, A. E. (Eds.), Advances

in Applied Microbiology. Academic Press. Inc., New York 1981,

pp. 85123.

[22] FAO/WHO. Joint FAO/WHO Expert Consultation on Eval-

uation of Health and Nutritional Properties of Probiotics inFood Including Powder Milk with Live Lactic Acid Bacteria.

2001. Cordoba, Argentina, 14 October, 2001. Available from:

http://www.fao.org/es/esn/food/foodandfoodprobioen.stm.

[23] Hempel, S., Newberry, S., Ruelaz, A., Wang, Z. et al., Safety

of probiotics used to reduce risk and prevent or tread dis-

ease: State of the research. Evidence Report/Technology As-

sessment No. HHSA290-2007-10062-I. AHRQ Rockville, MD:

Agence for Healthcare Research and Quality. Available from:

http://www.ahrg.gov/clinic/tp/probioctp.htm

[24] Servin, A. L., Antagonistic activities of lactobacilli and bifi-

dobacteria against microbial pathogens. FEMS Microbiol. Rev.

2004, 28, 405440.

[25] Maisnier-Patin, S., Deschamps, N., Tatini, S., Richard, J., In-hibition ofListeria monocytogenesin Camembert cheese made

with a nisin-producing starter. Lait1992, 72, 249263.

[26] Villani, F., Salzano, G.,Sorrentino, E.,Pepe, O.et al., Enterocin

226 NWC, a bacteriocin produced byEnterococcus faecalis226,

active against Listeria monocytogenes. J. Appl. Bacteriol. 1993,

74, 380387.

[27] Zottola, E. A.,Yezzi, T. L.,Ajao,D. B., Roberts, R. F., Utilization

of cheddar cheese containing nisin as an antimicrobial agent

in other foods. Int. J. Food Microbiol. 1994, 24, 227238.

[28] Ryan, M. P., Rea, M. C., Hill, C., Ross, R. P., An application

in cheddar cheese manufacture for a strain ofLactococcal lactis

producing a novel broad-spectrum bacteriocin, lacticin 3147.

Appl. Environ. Microbiol. 1996, 62, 612619.

[29] Nunez, M.,Rodriguez, J.L., Garcia, E.,Gaya, P. et al.,InhibitionofListeria monocytogenesby enterocin 4 during the manufac-

ture andripening of Manchegocheese.J. Appl. Microbiol. 1997,

83, 671677.

[30] Morgan, S., Ross, R. P., Hill, C., Increasing starter cell lysis in

cheddar cheese usinga bacteriocin-producingadjunct.J. Dairy

Sci. 1997, 80, 110.

[31] Garde, S., Gaya, P., Medina, M., Nunez, M., Acceleration of

flavour formation in cheese by a bacteriocin-producingadjunct

lactic culture. Biotechnol. Lett. 1997, 10, 10111014.

[32] Buyong, N., Kok, J., Luchansky, J. B., Use of a genetically en-

hanced, pediocin-producing starter culture, Lactococcal lactis

subsp. lactisMM217, to control Listeria monocytogenesin ched-

dar cheese. Appl. Environ. Microbiol. 1998, 64, 48424845.

[33] McAuliffe, O., Hill, C., Ross, R. P., Inhibition ofListeria mono-

cytogenesin cottage cheese manufactured with a lacticin 3147-

producing starter culture. J. Appl. Microbiol. 1999, 86, 251

256.

[34] Fenelon, M. A., Ryan, M. P., Rea, M. C., Guinee, T. P.

et al., Elevated temperature ripening of reduced fat cheddar

made with or without lacticin 3147-producing stsarter culture.

J. Dairy Sci . 1999, 82, 1022.

[35] Bouksaim, M., Lacroix, C., Audet, P., Simard, R. E., Effects

of mixed starter composition on nisin Z production by Lac-

tococcal lactis subsp. lactis biovar. diacetylactis UL 719 during

productionand ripening of goudacheese. Int.J. Food Microbiol.

2000, 59, 141156.

[36] Ryan, M. P., Ross, R. P., Hill, C., Strategy for manipulation of

cheese flora using combinations of lacticin 3147-producing

andresistant cultures. Appl. Environ. Microbiol. 2001, 67,

26992704.

[37] Martin-Cuesta, M. C., Requena, T., Pelaez, C., Use of a

bacteriocin-producing transconjugant as starter in accelera-tion of cheese ripening. Int. J. Food Microbiol. 2001, 70, 7988.

[38] Oumer, A., Gaya, P., Fernandez-Garcia, E., Mariaca, R. et al.,

Proteolysis and formation of volatile compounds in cheese

manufactured with a bacteriocin-producing adjunct culture.

J. Dairy Res. 2001, 68, 117129.

[39] Benech, R. O., Kheadr, E. E., Laridi, R., Lacroix, C. et al.,

Inhibition ofListeria innocuain cheddar cheese by addition of

nisin Z in liposomes or by in situ production in mixed culture.


[40] Sarantinopoulos, P., Leroy, F., Leontopoulou, E., Georgalaki,

M. D. et al., Bacteriocin production by Enterococcus faecium

FAIR-E 198in viewof itsapplicationas adjunctstarterin Greek

feta cheese making. Int. J. Food Microbiol. 2002, 72, 125136.[41] Morgan, S. M., O`Sullivan, L., Ross, R. P., Hill, C., The design

of a three strainstarter systemfor Cheddar cheesemanufacture

exploiting bacteriocin-induced starter lysis. Int. Dairy J. 2002,

12, 985993.

[42] O`Sullivan, L., Morgan, S. M., Ross, R. P., Hill, C., Elevated

enzyme release from lactococcal starter cultures on exposure

to the lantibiotic lacticin 481, produced by Lactococcus lactis

DPC 5552. J. Dairy Sci. 2002, 85, 21302140.

[43] O`Sullivan, L., Ross, R. P., Hill,C., A lacticin 481-producing ad-

junct culture increases starter lysis while inhibiting nonstarter

lactic acid bacteria proliferation. J. Appl. Microbiol. 2003, 95,

12351241.

[44] Rilla, N., Martinez, B., Deldago, T., Rodriguez, A., Inhibition

of Clostridium tyrobutyricum in Vidiago cheese by Lactococ-cus lactis ssp. lactis IPLA 729, a nisin Z producer. Int. J. Food

Microbiol. 2003, 85, 2333.

[45] Avila, M., Garde, S., Gaya, P., Medina, M. et al., Influence of a

bacteriocin-producing lacticculture on proteolysis and texture

of Hispanico cheese. Int. Dairy J. 2005, 15, 145153.

[46] Rodriguez, E., Calzada, J., Arques, J. L., Rodriguez, J. M. et al.,

Antimicrobial activity of pediocin-producing Lactococcus lac-

tis on Listeria monocytogenes, Staphylococcus aureus and Es-

cherichia coliO157:H7 in cheese. Int. Dairy J. 2005, 15, 5157.

[47] Bogovic Matijasic, B., Koman Rajsp, M., Perko, B., Rogelj, I.,

Inhibition of Clostridium tyrobutyricum in cheese by Lacto-

bacillus gasseri. Int. Dairy J. 2007, 17, 157166.

[48] Davies, E. A., Bevis, H. E., Delves-Broughton, J., The use of the

bacteriocin, nisin, as a preservative in ricotta-type cheeses to

control the food-borne pathogen Listeria monocytogenes. Lett.

Appl. Microbiol. 1997, 24, 343346.

[49] Aktypis, A., Kalantzopoulos, G., Huis in t` Veld, J. H. J., Ten

Brink, B., Purification and characterization of thermophilin

T, a novel bacteriocin produced byStreptococcus thermophilus

ACA-DC 0040. J. Appl. Microbiol. 1998, 84, 568576.

[50] Scannell, A. G. M., Hill, C.,Ross,R. P., Marx, S. et al., Develop-

ment of bioactive food packaging materials using immobilised

bacteriocins lacticin 3147 and nisaplin. Int. J. Food Microbiol.

2000, 60, 241249.



13/14


[51] Boussouel,N., Mathieu,F.,Revol-Junelles, A. M.,Milliere,J. B.,

Effects of combinations of lactoperoxidasesystem andnisin on

the behaviour ofListeria monocytogenesATCC 15313 in skim

milk. Int. J. Food Microbiol. 2000, 61, 169175.

[52] Mansour, M., Milliere, J. B., An inhibitory synergistic effect

of a nisin-monolaurin combination on Bacillusspp. vegetative

cells in milk. Food Microbiol. 2001, 18, 8794.

[53] Aktypis, A., Kalantzopoulos, G., Purification and character-ization of thermophilin ST-1, a novel bacteriocin produced

by Streptococcus thermophilus ACA-DC 0001. Lait 2003, 83,

365378.

[54] Franklin,N. B.,Cookey,K. D.,Getty,K. J., Inhibition ofListeria

monocytogeneson thesurfaceof individually packagedhot dogs

with a packaging film coating containing nisin. J. Food Prot.

2004, 67, 480485.

[55] Gilbreth, S., Somkuti, G. A., Thermophilin 110: a bacteriocin

ofStreptococcus thermophilusST 110. Curr. Microbiol. 2005, 51,

175182.

[56] Guerra, N. P., Macias, C. L., Agrasar, A. T., Castro, L. P., Devel-

opment of a bioactive packaging cellophane using nisaplin as

biopreservative agent. Lett. Appl. Microbiol. 2005, 40, 106110.[57] Mauriello, G., De Luca, E., La Storia, A., Villani, F. et al., An-

timicrobial activity of a nisin-activated plastic film for food

packaging. Lett. Appl. Microbiol. 2005, 41, 464469.

[58] Sobrino-Lopez, A., Martin Belloso, O., Enhancing inactiva-

tion ofStaphilococcus aureusin skim milk by combining high-

intensity pulsed electric fields and nisin. J. Food Prot. 2006, 69,

345353.

[59] Nes, I. F., Diep, D. B., Havarstein, L. S., Brurberg, M. B. et al.,

Biosynthesis of bacteriocins in lactic acid bacteria. Antonie van

Leeuwnhoek1996, 70, 113128.

[60] Holo, H., Nes, I. F., Class II antimicrobial peptides from lactic

acid bacteria. Biopolymers2000, 55, 5061.

[61] McAuliffe, O., Ross, R. P., Hill, C., Lantibiotics: structure,

biosynthesis and mode of action. FEMS Microbiol. Rev. 2001,25, 285308.

[62] Riley, M. A., Wertz, J. E., Bacteriocin diversity: ecological and

evolutionary perspectives. Biochimie2002, 84, 357364.

[63] Yamamoto, Y., Togawa, Y., Shimosaka, M., Okazaki, M., Pu-

rification and characterization of a novel bacteriocin produced

byEnterococcus faecalisstrain RJ-11. Appl. Environ. Microbiol.

2003, 69, 57465753.

[64] Rodriguez, J. M., Martinez, M. I., Horn, N., Dodd, H. M.,

Heterologousproduction of bacteriocins by lacticacid bacteria.

Int. J. Food Microbiol. 2003, 80, 101116.

[65] Sanchez-Hidalgo, M., Maqueda, M., Galvez, A., Abriouel, H.

et al., The genes coding for enterocin EJ 97 production by

Enterococcus faecalisEJ 97 are located on a conjugative plasmid.


[66] Kawai, Y., Ishii, Y., Arakawa, K., Uemura, K. et al., Structural

and functional differences in two cyclic bacteriocins with the

same sequences produced by lactobacilli. Appl. Environ. Micro-

biol. 2004, 70, 29062911.

[67] Fujita, K., Ichimasa, S., Zendo, T., Koga, S. et al., Structural

analysis and characterization of lacticin Q, a novel bacteri-

ocin belonging to a new family of unmodified bacteriocins

of Gram-positive bacteria. Appl. Environ. Microbiol. 2007, 73,

28712877.

[68] Wirawan, R. E., Swanson, K. M., Kleffmann, T., Jack, R. W.

et al., Uberolysin: a novel cyclic bacteriocin produced byStrep-

tococcus uberis. Microbiology2007, 153, 16191630.

[69] Abee, T., Krockel,L., Hill,C., Bacteriocins: Modes of actionand

potential in food preservation and control of food poisoning.

Int. J. Food. Microbiol. 1995, 28, 169185.

[70] Moll, G. N., Konings, W. N., Driessen, A. J. M., Bacteriocins:

mechanisms of membrane insertion and pore formation. An-tonie Van Leeuwnhoek1999, 76, 185198.

[71] Moll, G., Ubbink-Kok, T., Hildeng-Hauge, H., Nissen-Meyer,

J. et al., Lactococcin G is a potassium ion-conducting, two-

component bacteriocin. J. Bacteriol. 1996, 178, 600605.

[72] Nissen-Meyer, J., Holo, H., Havarstein, L. S., Sletten, K. et al.,

A novel lactococcal bacteriocin whose activity on the comple-

mentary action of two peptides. J. Bacteriol. 1992, 174, 5686

5692.

[73] Fremaux, C., Ahn, C., Klaenhammer, T. R., Molecular analysis

of the lacticin operon. Appl. Environ. Microbiol. 1993, 59, 3906

3915.

[74] Sahl, H. G.,Pore formation in bacterialmembranesby cationic

lantibiotics, in: Jung, G., Sahl, H.G. (Eds.), Nisin and NovelLantibiotics, ESCOM Science, Leiden 1991, pp. 347358.

[75] Matsusaki,H., Sonomoto, K., Ishizaki,A., Some characteristics

of nisin Z, a peptide antibiotic produced byLactococcus lactis

IO-1. Food Sci. Technol. Int. 1998, 4, 290294.

[76] Bard, S., Abdel Karem, H., El-Hadedy, D., Characterization of

nisin produced byLactococcus lactis. Int. J. Agric. Biol. 2007, 3,

499503.

[77] Ryan, M. P., Rea, M. C., Hill, C., Ross, R. P., An application in

cheddar cheese manufacture for a strain of Lactococcus lactis

producing a novel broad-spectrum bacteriocin, lacticin 3147.


[78] Morgan, S. M., Ross, R. P., Beresford, T., Hill, C., Combina-

tion of hydrostatic pressure and lacticin 3147 causes increased

killing ofStaphylococcusand Listeria. J. Appl. Microbiol. 2000,88, 414420.

[79] Cunniffi, A., Isolation and characterization of two bacteriocino-

genic Lactococcal strains. MSc Thesis, National University of

Ireland, Cork, Irland 1998.

[80] Ennahar, S., Aoude-Werner, D., Assobhel, O., Hasselmann, C.,

Antilisterial activity of enterocin 81, a bacteriocin produced

byEnterococcus faecium WHE 81 isolated from cheese. J. Appl.

Microbiol. 1998, 85, 521526.

[81] Ananou, S., Galvez, A., Martnez-Bueno, M., Maqueda, M.

et al., Synergistic effectof enterocin AS-48 in combination with

outer membrane permeabilizing treatment against Escherichia

coliO157:H7. J. Appl. Microbiol. 2005, 99, 13641372.

[82] Tulini, F. L., Gomes, B. C., De Martinis, E. C. P., Partial purifi-

cation and characterization of a bacteriocin produced by En-

terococcus faecium 130 isolated from mozzarella cheese. Cienc.

Tecnol. Aliment. 2011, 31, 155159.

[83] Rodrguez, J. M., Martnez, M. I., Kok, J., Pediocin PA-1, a

wide-spectrum bacteriocin from lactic acid bacteria. Crit. Rev.

Food Sci. Nutr. 2002, 42, 91121.

[84] Abo-Amer, A. E., Characterization of a bacteriocin-like in-

hibitory substance produced by Lactobacillus plantarum iso-

lated from Egyptian home-made yogurt. Sci. Asia, 2007, 33,

313319.



14/14


[85] Hernandez,D., Cardell, E.,Zarate, V., Antimicrobial activity of

lactic acid bacteria isolated from Tenerife cheese: initial char-

acterization of plantaricin TF711, a bacteriocin-like substance

produced byLactobacillus plantarum TF711.J. Appl. Microbiol.

2005, 99, 7784.

[86] Chumchalova, J., Stiles, J., Josephsen, J., Plockova, M., Char-

acterization and purification of acidocin CH5, a bacteriocin

produced byLactobacillus acidophilusCH5. J. Appl. Microbiol.2004, 96, 10821089.

[87] Miteva, V., Ivanova, I., Budakov, I., Pantev, A. et al., Detec-

tion and characterization of a novel antibacterial substance

produced by a Lactobacillus delbrueckii strain 1043. J. Appl.

Microbiol. 1998, 85, 603614.

[88] Tambekar,D.H.,Bhutada,S.A.,Studiesonantimicrobialactiv-

ity and characterictics of bacteriocins produced byLactobacil-

lusstrains isolated from milk of domestic animals. Internet J.

Microbiol., 2010, 8. Available from: http://www.ispub.com.

[89] Simova, E., Beshkova, D., Najdenski, H., Frengova, G. et al.,

Antimicrobial-producing lactic acid bacteria isolated from tra-

ditional Bulgarian milk products: inhibitory properties and in

situ bacteriocinogenic activity. IUFOST, 13th World Congressof Food Science and Technology Food is Life, Nantes, France,

1721 September, 2006, 907908.

[90] Casla, D., Requena, T., Gomez, R., Antimicrobial activity of

lactic acid bacteria isolated from goats milk and artisanal

cheeses: characteristics of a bacteriocin produced by Lacto-

bacillus curvatus IFPL105. J. Appl. Bacteriol. 1996, 81, 35

41.

[91] Villani, F., Pepe, O., Mauriello, G., Salzano, G. et al., Antilis-

terial activity of thermophilin 347, a bacteriocin produced by

Streptococcus thermophilus. Int.J.FoodMicrobiol. 1995, 25,179

190.

[92] Ivanova, I., Miteva, V., Stefanova, T. S., Pantev, A. et al., Char-

acterization of bacteriocin produced by Streptococcus ther-

mophilus81. Int. J. Food Microbiol. 1998, 42, 147158.

[93] Powell, I. B., Broome, M. C., Limsowtin, G. K. Y., Starter cul-

tures: general aspects, in: Roginski, H., Fuquay, J. W., Fox, P. F.

(Eds.), Encyclopedia of Dairy Science. Academic Press, London2002, pp. 261268.

[94] El Soda, M., Madkor,S. A.,Tong, P. S.,Adjunct cultures:recent

developments and potential significanceto the cheese industry.

J. Dairy Sci. 2000, 83, 609619.

[95] Folkertsma, B., Fox, P. F., McSweeney, P. L. H., Accelerated

ripening of cheese at elevated temperatures. Int. Dairy J. 1996,

6, 11171124.

[96] Reviriego, C., Fernandez, A., Horn, N., Rodriguez, E. et al.,

Production of pediocin PA-1, and coproduction of nisin A and

pediocin PA-1, by wildLactococcuslactisstrains of dairy origin.

Int. Dairy J. 2005, 15, 4549.

[97] Avonts,A., Van Uytven, E.,De Vuyst,L., Cell growthand bacte-

riocin production of probiotic Lactobacillusstrains in differentmedia. Int. Dairy J. 2004, 14, 947977.

[98] Giraffa, G., Neviani, E., Torri Tarelli, G. Antilisterial activity of

enterococci in a model predicting the temperature evolution

of Taleggio, an Italian soft cheese. J. Dairy Sci. 1994, 77, 1176

1182.

[99] Garde, S., Avila, M., Medina, M., Nunez, M., Proteolysis of

Hispanico cheese manufactured using lacticin 481-producing

Lactococcus lactis ssp. lactis INIA 639. J. Dairy Sci. 2006, 89,

840849.


Bacteriocin of LAB Importance for Dairy Industry

Documents