Ci a ion: We ning, M.L.;
He nández-Alcán a a, A.M.; Ruiz,
M.J.; So o, L.P.; Dueñas, M.T.; López,
P.; F izzo, L.S. Biological Func ions o
Exopolysaccha ides om Lac ic Acid
Bac e ia and Thei Po en ial Bene i s
o Humans and Fa med Animals.
Foods 2022,11, 1284. h ps://doi.o g/
10.3390/ oods11091284
Academic Edi o : Salam A. Ib ahim
Recei ed: 31 Ma ch 2022
Accep ed: 25 Ap il 2022
Published: 28 Ap il 2022
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Licensee MDPI, Basel, Swi ze land.
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A ibu ion (CC BY) license (h ps://
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oods
Re iew
Biological Func ions o Exopolysaccha ides om Lac ic Acid
Bac e ia and Thei Po en ial Bene i s o Humans and
Fa med Animals
Ma ía Lau a We ning 1,*, Annel M. He nández-Alcán a a 2, Ma ía Julia Ruiz 1,3 , Lo ena Paola So o 1,4,
Ma ía Te esa Dueñas 5, Paloma López 2and Lau eano Sebas ián F izzo 1,4
1Labo a o y o Food Analysis “Rodol o Osca DALLA SANTINA”, Ins i u e o Ve e ina y Science
(ICiVe Li o al), Na ional Uni e si y o he Li o al-Na ional, Council o Scien i ic and Technical
Resea ch (UNL/CONICET), Espe anza 3080, SF, A gen ina; j [email p o ec ed] (M.J.R.);
[email p o ec ed] (L.P.S.); l izzo@ c .unl.edu.a (L.S.F.)
2Depa men o Mic oo ganisms and Plan Bio echnology, Ma ga i a Salas Cen e o Biological
Resea ch (CIB)-Consejo Supe io de In es igaciones Cien í icas (CSIC), 28040 Mad id, Spain;
[email p o ec ed] (A.M.H.-A.); [email p o ec ed] (P.L.)
3Depa men o Animal Heal h and P e en i e Medicine, Facul y o Ve e ina y Sciences, Na ional Uni e si y
o he Cen e o he P o ince o Buenos Ai es, Buenos Ai es 7000, A gen ina
4Depa men o Public Heal h, Facul y o Ve e ina y Science, Li o al Na ional Uni e si y,
Espe anza 3038, A gen ina
5Depa men o Applied Chemis y, Facul y o Chemis y, Uni e si y o he Basque Coun y (UPV/EHU),
20018 San Sebas ián, Spain; [email p o ec ed]
*Co espondence: [email p o ec ed]; Tel.: +54-3496420639 (ex . 421037)
Abs ac :
Lac ic acid bac e ia (LAB) syn hesize exopolysaccha ides (EPS), which a e s uc u ally
di e se biopolyme s wi h a b oad ange o echnological p ope ies and bioac i i ies. The e is
scien i ic e idence ha hese polyme s ha e heal h-p omo ing p ope ies. Mos comme cialized
p obio ic mic oo ganisms o consump ion by humans and a med animals a e LAB and some o
hem a e EPS-p oduce s indica ing ha some o hei bene icial p ope ies could be due o hese
polyme s. P obio ic LAB a e cu en ly used o imp o e human heal h and o he p e en ion and
ea men o speci ic pa hologic condi ions. They a e also used in ood-p oducing animal husband y,
mainly due o hei abili ies o p omo e g ow h and inhibi pa hogens ia di e en mechanisms,
among which he p oduc ion o EPS could be in ol ed. Thus, he aim o his e iew is o discuss he
cu en knowledge o he cha ac e is ics, usage and biological ole o EPS om LAB, as well as hei
pos bio ic ac ion in humans and animals, and o p edic he u u e con ibu ion ha hey could ha e
on he die o ood animals o imp o e p oduc i i y, animal heal h s a us and impac on public heal h.
Keywo ds: exopolysaccha ides; EPS; lac ic acid bac e ia; LAB; p obio ics; ood-p oducing animals
1. In oduc ion
Lac ic acid bac e ia (LAB) a e G am-posi i e, anae obic ae o ole an , non-spo e-
o ming bac e ia wi h od o coccus shape. They cons i u e a g oup o bac e ia commonly
ound in dai y ( e men ed), mea and ege able p oduc s, as well as in he gas oin es inal
and u ogeni al ac s o humans and animals, on skin, and in soil and wa e [
1
,
2
]. LAB
a e used as s a e cul u es o e men a ions and as p obio ics in unc ional oods, and
he p oduc ion o compounds such as nu aceu icals [
3
,
4
]. Many LAB can syn hesize
ex acellula polysaccha ides (EPS) o high molecula weigh . These polyme s a e s ain
speci ic and display g ea s uc u al di e si y. Some EPS emain in ima ely a ached o he
su ace o he bac e ia o ming a capsule [
5
]. Howe e , in his e iew, he e m EPS will
e e o hose non-capsula polysaccha ides ha , al hough hey may be loosely associa ed
wi h he bac e ial su ace, a e la gely eleased in o he ex e nal en i onmen .
Foods 2022,11, 1284. h ps://doi.o g/10.3390/ oods11091284 h ps://www.mdpi.com/jou nal/ oods
Foods 2022,11, 1284 2 o 34
EPS a e homopolysaccha ides (HoPS) o he e opolysaccha ides (HePS), depending on
whe he hei main chain is composed o one o a ious monome s and hei mechanisms
o syn hesis a e, in gene al, ex a- o in a-cellula , espec i ely [6].
EPS-p oducing LAB belong o a ious gene a dis ibu ed in mul iple habi a s. The
mos p ominen HePS-p oducing LAB a e Lac obacillus,Lac ococcus,S ep ococcus and En e-
ococcus s ains equen ly isola ed om e men ed dai y p oduc s and he human gas oin-
es inal ac (GIT) and eces, whe eas he majo i y o HoPS-p oducing LAB a e Lac obacillus,
Leuconos oc,Pediococcus,S ep ococcus, and Weissella s ains isola ed om animal GIT, eg-
e ables and e men ed be e ages [
7
]. The wide dis ibu ion o EPS p oduc ion indica es
ha i mus con e some compe i i e ad an age in he ecological niches o hese bac e ia.
Indeed, some possible physiological oles p oposed o LAB-EPS a e he p o ec ion o he
bac e ia om s ess condi ions, including en i onmen al pH, osmo ic s ess o desicca ion,
and om bac e iophages, an ibio ics, and lysozymes, enabling LAB pe sis ence in a ious
niches including bio ilm o ma ion [
8
,
9
]. In gene al, syn hesis o EPS is no in ol ed in
he gene a ion o ene gy, nei he a e EPS used as a ca bon sou ce by he p oducing mi-
c oo ganism [
10
]. Ne e heless, syn hesis o he HoPS dex an, by Weissella and Leuconos oc
s ains, as well as lac obacilli, is accompanied by he hyd olysis o suc ose gene a ing
monosaccha ides which a e used by hese LAB as a ca bon sou ce [11].
Se e al applica ions o EPS om LAB ha e been p oposed. Wi h he excep ion o
HoPS dex an, un il now, only he in si u applica ion o EPS-p oducing LAB has been
economically iable, o example, as s a e cul u es, ins ead o pu i ied EPS due o he
insu icien le els no mally p oduced, as well as p oduc ion cos s. Pa icula ly in he
e men ed ood indus y, EPS p oduc ion by LAB con ibu es o imp o ing he o ganolep ic
quali y, senso y and heological p ope ies, as well as s abili y, o he inal p oduc s. Mo e-
o e , by hei addi ion o hese ood p oduc s, EPS can be conside ed as unc ional pos bio ic
ing edien s due o hei pos ula ed human heal h bene i s, such as immuno-modula ion,
an i-oxida i e, an i-in lamma o y, an i-mic obial, an i- umo al, o choles e ol-lowe ing
p ope ies, o as mic obiome modula o s [
12
–
14
]. Indeed, many p obio ic bac e ia (o
bac e ia wi h po en ial as such), a e p oduce s o a wide ange o EPS and he scien i ic
e idence sugges s ha some o he heal h e ec s a ibu ed o p obio ic LAB may be due o
hei p oduc ion o he polyme s. Thus, bac e ial esis ance o gas oin es inal s ess and he
pe sis ence o he bac e ia in he gu ecosys em a e in luenced by hei EPS p oduc ion [
9
].
The p esen wo k includes an upda ed e iew o gene al issues ela ed o LAB-EPS
including i s classi ica ion, biosyn hesis and p oduc ion and especially i s cu en and
po en ial applica ions. Howe e , he main objec i e o his e iew is o de ail he cu en
knowledge o EPS p oduc ion by p obio ic LAB. The ole o LAB-EPS is e iewed in de ail,
bo h in e ms o he p obio ic p ope ies and he biological ac i i ies a ibu ed o hei
p oducing bac e ia and whose e ec s can be bo h local and sys emic in he hos . Finally,
he au ho s conside he po en ially bene icial u u e con ibu ion hey could make by
inclusion in he die o a med animals o he imp o emen o p oduc i i y, sani a ion and
public heal h.
2. S uc u e, P oduc ion and Pu i ica ion o EPS
2.1. S uc u e
LAB p oduce a wide di e si y o EPS as de ined by hei monome composi ion, molec-
ula mass and s uc u e. Acco ding o hei monosaccha ide composi ion and biosyn he ic
pa hway, hey a e classi ied as HePS o HoPS. HePS possess a iable molecula masses
(gene ally up o 10
6
Da) wi h wo o mo e di e en ypes o monosaccha ides o ganized
in epea ing uni s wi h a a iable numbe om i- o oc a-saccha ides. The monome s,
equen ly D-glucose, D-galac ose, and L- hamnose a e usually joined by
β
-(1,4) o
β
-(1,3)
and
α
-(1,2) o
α
-(1,6) linkages, bu o he componen s, such as L- ucose, D-mannose, N-
ace ylmonosaccha ides, D-glucu onic acid o glyce ol may also be p esen [
6
]. In addi ion,
hey can be modi ied wi h py u a e and phospha e. Ou o he 81 epo ed s uc u es, a
monosaccha ide occu ence in 55 unique epea ing uni s has been desc ibed; among hem,
Foods 2022,11, 1284 3 o 34
40 co espond o Lac obacillus, 11 o S ep ococcus and 4 o Lac ococcus gene a [
15
]. The la ge
a ia ion in composi ion, linkage ype and b anching pa e ns leads o a la ge di e si y in
he polyme s uc u es and sizes, esul ing in di e se echnological unc ions.
2.2. P oduc ion
The HePS a e syn he ized by in acellula glycosyl ans e ases using suga nucleo ides
as subs a es. These enzymes a e encoded by genes loca ed in he eps gene clus e , oge he
wi h o he genes encoding p o eins in ol ed in chain-leng h, polyme iza ion and expo
o he EPS, as well as in he egula ion o gene exp ession [
5
,
6
]. Al hough he exac
mechanisms a e no ully unde s ood, he ole o he molecula de e minan s o LAB-EPS
biosyn hesis has been ecen ly e iewed [7,9,15].
Wi h espec o HoPS, hese polysaccha ides a e widely p oduced by LAB, being
epo ed in species belonging o Weissella,S ep ococcus,Pediococcus,Oenococcus,Lac obacillus
and Leuconos oc gene a; hei p oduc ion has been ex ensi ely e iewed in [
8
,
12
]. In
gene al, hese polyme s display high molecula masses (up o 10
8
Da) and a e composed
o glucose (glucans), uc ose ( uc ans) o galac ose (polygalac ans). These polyme s a e
subdi ided based on he ype o glycosidic linkage, and he ype and deg ee o b anching.
Glucans include
α
-glucans (e.g., dex an, mu an, al e nan and eu e an) and
β
-glucans,
whe eas he e a e wo classes o uc ans, le an- ype and inulin- ype, bo h being
β
- uc ans.
Ex acellula glycansuc ases, encoded by a single gene, ca alyse he syn hesis o he majo i y
o hese HoPS using suc ose as a subs a e.
Howe e , o he mechanisms o glucan biosyn hesis ha e been epo ed. Thus,
β
-
glucans a e syn hesized by a single ansmemb ane glycosyl ans e ase ha u ilizes UDP-
glucose as he dono subs a e [
16
]. Fu he mo e, Maye e al. [
17
] ha e ecen ly epo ed
ha a pu a i e bac op enol glycosyl ans e ase and lippase a e essen ial o b anched
dex an HoPS biosyn hesis in L. johnsonii F19785.
Bo h, he yield and molecula mass o he EPS p oduced by LAB can be a ec ed
by in insic, as well as en i onmen al ac o s, including medium composi ion, g ow h
condi ions and incuba ion ime. HePS a e syn hesized in low amoun s (20 o 600 mg/L),
al hough, unde op imal cul u e condi ions, a ew s ains p oduce highe amoun s, e.g.,
S ep ococcus he mophilus ASCC 1275 and Lac icaseibacillus hamnosus RW-9595M ( o me ly
Lac obacillus hamnosus) p oducing 1 g/L, and 2.7 g/L, espec i ely. Howe e , p oduc ion
o HoPS is high, wi h some s ains syn hesizing e en 10 g/L, e.g., Limosilac obacillus eu e i
Lb121 ( o me ly Lac obacillus eu e i), which p oduces an
α
-glucan and a
β
- uc an. The
di e ence in he p oduc ion o bo h classes o polyme s may be because he syn hesis o he
HePS, compa ed o ha o he HoPS, is a mo e complex mechanism ela ed o he cen al
me abolism o he bac e ium. In addi ion, he isola ion and pu i ica ion p ocedu es can
lead o signi ican di e ences in he polysaccha ide concen a ion measu ed [6,15].
Fo he echnological applica ion o EPS om LAB, an op imal and su icien p oduc-
ion, bo h in si u and ex si u, is impe a i e, no only o imp o e hei yield, bu also o
ob ain a pa icula unc ionali y. In his espec , me abolic enginee ing could imp o e EPS
p oduc ion, al hough i migh be o limi ed alue due o he inhe en complexi y o he
syn he ic mechanisms, especially in he case o HePS [
18
]. Ne e heless, an example o
his was he inding ha NADH oxidase o e exp ession had a signi ican e ec on EPS
p oduc ion in Lac icaseibacillus casei LC2W ( o me ly Lac obacillus casei) [18].
Ano he s a egy is he op imiza ion o he p oduc ion condi ions, as well as he
egula ion mechanisms. En i onmen al s ess can imp o e he p oduc ion o EPS by LAB
and p oduce cus omized EPS wi h desi ed unc ionali y. Fo example, EPS syn hesis by L.
hel e icus ATCC 15807 is s imula ed unde acidic pH s ess and inhibi ed unde sodium
chlo ide osmo ic s ess [
19
]. Mo eo e , in L. hel e icus 6E8, a high suga concen a ions,
he e was a shi om bac e ial g ow h o EPS syn hesis and sec e ion [20].
The s udy o he condi ions and mechanisms o exp ession egula ion o dex an p o-
duc ion in Leuconos oc lac is AV1n e ealed ha his s ain syn hesizes he HoPS e icien ly
Foods 2022,11, 1284 4 o 34
a low empe a u e; he e o e, his LAB could be a good candida e o he in si u p oduc ion
o he polyme du ing he manu ac u e o some unc ional oods (e.g., ke i ) [11].
2.3. Pu i ica ion
Bo h classes o polysaccha ides can o en be pu i ied by simila p ocedu es. Howe e ,
o achie e he bes possible yield, i is necessa y o es ablish a combina ion o condi ions
and p ocedu es o each case [
21
]. Labo a o y me hods o he p oduc ion, isola ion, pu i i-
ca ion and quan i ica ion o EPS we e ecen ly e iewed p o iding a de ailed desc ip ion
o his opic [
18
,
22
]. B ie ly, EPS can ypically be eco e ed om bac e ial cul u e supe -
na an s using a ia ions o a gene al me hod, including p ecipi a ion om he b o h, and
pu i ica ion o he polyme om he p ecipi a e. Depending on he medium, he i s s ep
is o en p o ein elimina ion using ichlo oace ic acid o p o eases. A e cen i uga ion, he
biopolyme s a e eco e ed om he g ow h medium by p ecipi a ion wi h cold e hanol
o ace one. The EPS is hen dissol ed in dis illed wa e and dialyzed in a bag wi h a
12–14-kDa cu -o o elimina e low-molecula -mass ca bohyd a es, and, subsequen ly, he
EPS a e lyophilized. Addi ional ch oma og aphic pu i ica ion s eps a e usually equi ed i
he s uc u e o he EPS is o be cha ac e ized and po en ial biological applica ions a e o
be explo ed.
The s uc u es o EPS ha e been ex ensi ely in es iga ed; gene ally, hei s udy
includes de e mina ion o molecula weigh by gel pe mea ion ch oma og aphy and
asymme ic ield- low ac iona ion (AF4) o by size exclusion ch oma og aphy coupled o
mul iple-angle lase ligh sca e ing, he composi ion o monosaccha ides by liquid-gas
ch oma og aphy ollowed by mass spec ome y, and he monosaccha ide pa e n by a
combina ion o me hyla ion and nuclea magne ic esonance analysis. Finally, he mi-
c os uc u es o he EPS a e obse ed by scanning elec on mic oscopy o a omic o ce
mic oscopy [7].
The chemical s uc u es and molecula masses a e p ope ies ha ha e been co ela ed
wi h he echnological cha ac e is ics and biological unc ions o he EPS in di e se appli-
ca ions. A comp ehensi e e iew o he ela ionship be ween he s uc u e and unc ion
o EPS om LAB and bi idobac e ia has ecen ly been epo ed [
7
,
23
]. Howe e , in he
ollowing sec ions, he pos bio ic e ec s known o da e o EPS om LAB will be discussed,
in pa icula , as well as hei ole in he p obio ic p ope ies o he p oducing LAB.
3. Biological Func ions o he EPS
3.1. An imic obial E ec s
The an imic obial ac i i y o a ious EPS om LAB agains a wide a ie y o pa hogenic
mic obes has been in es iga ed using bo h
in i o
and
in i o
assays [
24
–
26
]. EPS can exe
hei an imic obial ac ion indi ec ly, ei he (i) h ough he s imula ion o he inna e and
adap i e immune esponse, o (ii) by p omo ing he g ow h and/o o ma ion o bio ilms
o o he bene icial commensal bac e ia o p obio ics. Al hough hese opics a e discussed
in he immunomodula o y e ec s and p ebio ic p ope ies sec ions, espec i ely, some ex-
amples o EPS ac ion agains pa hogens h ough bo h modes o ac ion a e desc ibed below.
Dex an p oduced by L. mesen e oides NTM048 s imula ed mucosal IgA sec e ion. Using an
in i o
assay, NTM048 EPS s imula ed T helpe (Th1) and (Th2) cells-media ed esponses,
as well as o al and an igen-speci ic IgA p oduc ion in splenocy es [
27
]. In addi ion, i
has been demons a ed
in i o
ha he HePS om L. e men um UCO-979C is pa ially
esponsible o inc eased esis ance agains Helicobac e pylo i in ec ion by modula ing he
gas ic inna e immune esponse [28].
The an imic obial ac i i y o LAB-EPS could also be explained
in i o
by: (i) he
p ebio ic e ec ha helps LAB in gu coloniza ion, (ii) he p o ec ion o commensal mic oo -
ganisms om he adap i e immune esponse in he hos , and (iii) he enhancemen o hei
compe i ion wi h pa hogenic bac e ia [
29
]. Al hough he e a e no ela ed epo s in LAB,
his was demons a ed o he HePS om Bi idobac e ium b e e UCC2003 [
30
] and HoPS
om Bacillus sub ilis HMNig-2 [31] in mu ine models.
Foods 2022,11, 1284 5 o 34
These polyme s can also di ec ly p omo e an imic obial ac i i y in he ollowing ways:
(i) inhibi ing he g ow h o pa hogenic mic oo ganisms; (ii) in e e ing wi h hei adhesion
o he in es inal epi helium; and (iii) by p e en ing o educing he o ma ion o bio ilms
by pa hogenic bac e ia. The an i-bio ilm p ope ies o LAB-EPS a e examined below; some
examples a e b ie ly men ioned. HePS p oducing L. hamnosus and HePS om L. plan a um
WLPL04 exhibi ed s ong inhibi ion agains bio ilm o ma ion by pa hogenic bac e ia,
including Salmonella en e ica se o a yphimu ium [
32
], as well as Pseudomonas ae uginosa,
Esche ichia coli O157:H7, and S aphylococcus au eus [
33
]. Mo eo e , he dis up i e e ec o
EPS p oduced by Loigolac obacillus co yni o mis NA-3 ( o me ly Lac obacillus co yni o mis) on
p e- o med S. yphimu ium and Bacillus ce eus bio ilms was he g ea es o hose es ed and
was epo ed o be 80 and 90%, espec i ely [
7
]. A possible mechanism o an imic obial
ac ion is ha he EPS o LAB in e ac wi h he signaling molecules in ol ed in he o ma ion
o bio ilms on he su ace o hese pa hogens, in e up ing hei o ma ion and hus exe ing
an an imic obial e ec [34].
Wi h espec o he abili y o EPS o inhibi pa hogenic mic oo ganisms, i was epo ed
ha HePS p oducing L. hamnosus isola ed om human b eas milk showed, in an
in i o
assay, s ong an i-bac e ial ac i i y agains pa hogenic E. coli and S. yphimu ium, [
32
]. Simi-
la ly, HePS p oduced by L. gasse i [
35
] and L. ke i ano aciens DN1 [
36
] ha e shown
in i o
an ibac e ial ac i i y agains se e al oodbo ne pa hogens, such as Lis e ia monocy ogenes
and Salmonella en e i idis. Mo eo e , in a ecen s udy, HePS ex ac ed om L. plajomi PW-7
( o me ly Lac obacillus plajomi) showed an ibac e ial ac i i y agains H. pylo i,S. au eus, and
E. coli [
37
]. Wi h espec o he po en ial inhibi o y mechanism o pa hogenic bac e ia by
EPS, i was p oposed ha his can be e ec ed by dis up ing he in eg i y o memb anes
and eleasing he con en s o soluble p o eins [
38
]. Indeed, he dis up ion o he cell mem-
b anes o some G am-posi i e and G am-nega i e pa hogenic bac e ia by EPS om L.
plajomi PW-7 was con i med by elec on mic oscopy [
37
]. Ano he mechanism, p oposed by
Salachna e al. [
39
], is ha he EPS may p omo e he accumula ion o seconda y me aboli es
in he g ow h media, which migh ad e sely a ec bo h G am-posi i e and G am-nega i e
pa hogens [39].
The in e ac ion o he bac e ial EPS ke i wi h bac e ial o euka yo ic cells, sugges s
ha he an imic obial ac ion o his EPS occu s ia he blockade o ecep o s o channels o
he ou e memb ane [40].
An i ungal ac i i y o LAB-EPS has been epo ed [
24
]. The ac i i y o EPS-p oducing
L. hamnosus GG, i s non-p oducing mu an as well as pu i ied EPS, e ealed ha GG EPS
migh be in ol ed in educing hyphal o ma ion and dec easing Candida adhesion h ough:
(i) co-agg ega ion, (ii) immunomodula ion o he hos epi helial cells, and (iii) compe i ion
o binding si es [41].
Fu he mo e, i has been epo ed ha EPS p omo ed an i i al ac i i ies by im-
munomodula ing na i e and adap i e esponse and by in e e ing wi h he adhesion
and /o ep oduc ion o i al pa icles (an i i al p ope ies linked wi h immune sys em
esponses a e discussed in he an i i al e ec s sec ion). On he o he hand, some s udies
sugges ha EPS can ac di ec ly agains a ious i al pa hogens [
24
]. Wi h espec o his
e ec o mode o ac ion, i was shown ha EPS 26a om Lac obacillus spp. obs uc ed
adeno i us ype-5 (HAdV-5) ep oduc ion [
42
] and EPS om L. plan a um LRCC5310 in e -
e ed wi h o a i us a achmen o cells
in i o
[
43
]. In addi ion, es s in young mice, wi h
EPS-LRCC5310, p oduced a educ ion in o a i us eplica ion in he in es ine and in he
du a ion o dia hea, wi h consequen educ ion in he eco e y ime o he mice [43].
The in e e ence ac i i y exe ed by LAB-EPS on pa hogens in he hos can be assessed
h ough he abili y o co-agg ega e and dec ease he accessibili y o pa hogens o he
in es inal epi helium. S ains o L. delb ueckii subsp. bulga icus can co-agg ega e wi h E.
coli [
44
]. This mode o ac ion has also been demons a ed by u iliza ion o in es inal cell line
models. L. pa aplan a um BGCG11, a p oduce o HePs, bu no i s isogenic non-p oduce
s ain, showed a p o ec i e e ec on mucous memb anes by hinde ing he con ac o E. coli,
L. monocy ogenes wi h epi helial cells HT29-MTX [
45
]. In ano he s udy, i was de e mined
Foods 2022,11, 1284 6 o 34
ha he HePS p oduced by he s ain L. pa acasei subsp. pa acasei BGSJ2-8 was in ol ed
in adhesion o epi helial in es inal cells and dec eased he E. coli associa ion o Caco-2
cells [
46
]. In addi ion, HePS-p oducing L. plan a um WLPL04 isola ed om human b eas
milk had a signi ican inhibi o y e ec on he adhesion o E. coli O157:H7 o human in es inal
epi helial cells [
47
]. A pos e io i pu i ied HePS-WLPL04 had a simila inhibi o y e ec on
he adhesion o E. coli O157:H7 o HT-29 cells in compe i ion, eplacemen , and inhibi ion
assays [
33
]. In addi ion, in an
in i o
s udy, L. johnsonii FI9785, a HePS and
α
-glucan
p oduce , was e ec i e in supp essing he coloniza ion and pe sis ence o Clos idium
pe ingens in poul y and educed coloniza ion by E. coli o he small in es ine [
48
,
49
].
Th ough hyd ophobici y and au oagg ega ion, EPS syn hesized by L. johnsonii FI9785
compe i i ely inhibi ed hese pa hogens [50].
EPS can also exe an an agonis ic e ec on he in lamma o y esponse caused by
pa hogenic mic oo ganisms in he in es inal epi helium h ough modula ion o he immune
sys em. Po cine in es inal epi helial cell lines ha e been used o de e mine he an agonis-
ic e ec s o EPS-p oducing s ains, such as L. delb ueckii TUA4408L, by a enua ing he
in lamma o y esponse induced by en e o oxigenic E. coli [51].
The an imic obial ac ion o EPS depends on hei composi ion and s uc u e. The
molecula weigh , composi ion, and cha ged g oups a e epo ed o be pa icula ly ela ed
o his ac i i y. Pu i ied EPS ac ions o high molecula size showed s onge an ibac e ial
e ec s agains G am-nega i e bac e ia, whe eas he opposi e end was obse ed o G am-
posi i e bac e ia [
52
]. Mo eo e , EPS composi ion is implica ed in he po en ial in e ac ion
wi h pa hogens. In his espec , analysis o he EPS-de icien mu an L. pa acasei subsp.
pa acasei BGSJ2-8, which p oduces an EPS wi h a di e en composi ion om ha o he
wild- ype s ain, indica ed ha he wild- ype EPS is essen ial o educe E. coli associa ion
wi h Caco-2 cells [
46
]. Mo eo e , in a s udy ha e alua ed he an ibac e ial e ec o L.
plajomi PW-7-EPS componen s, i was ound ha galac ose had a g ea e e ec on E. coli,
while glucu onic acid had a g ea e e ec on S. au eus and xylose had a s onge e ec on
H. pylo i [37].
Finally, subs i u ional modi ica ions o EPS, including sul ona ion, phospho yla ion
and ace yla ion ha e been epo ed o a ec hei an imic obial ac i i y. The sul a ion
o EPS leads o po en an i i al ac i i y. The modi ica ion was shown o be e ec i e in
inhibi ing i us-cell in e ac ion. In addi ion, his ac i i y was demons a ed, among o he s,
o hepa i is B, he pes and in luenza i uses [
53
]. Sul a ed EPS om L. plan a um ZDY2013
and S. he mophilus exhibi ed a g ea e an imic obial e ec agains a ious G am-posi i e
and ha m ul pa hogens han non-sul a ed compounds [54,55].
3.2. An i-Bio ilm P ope ies
Bio ilms a e ex acellula ma ices which adhe e o su aces, composed o a complex
o nucleic acids, p o eins, polysaccha ides and lipids. Many bac e ial species, including
pa hogenic bac e ia, become mo e esis an o ex acellula s ess condi ions by p oducing
bio ilms. In se e al pa hogenic mic oo ganisms, en i onmen al s ess can igge he
o ma ion o bio ilms, which inc ease adhesion and p o ec ion agains he hos esponse.
Consequen ly, bio ilms play an impo an ole in pa hogenesis [
56
]. Coloniza ion o he
chicken gas oin es inal ac and o iduc s by S. yphimu ium is la gely due o i s abili y o
adhe e and o o m bio ilms. In chicken epi helial cell lines (Hep-2), i has been shown ha
ce ain EPS con ibu e o bio ilm o ma ion [57].
The e is inc easing scien i ic e idence suppo ing he posi ion ha a ious EPS ex-
ac ed om LAB can educe o inhibi mic obial bio ilms and ha , he e o e, hey ha e
po en ial applica ion in he design o new s a egies o deal wi h bac e ial bio ilm-associa ed
in ec ions and ood sa e y issues [
24
]. I has been shown ha many EPS o lac obacilli
can in e ene in he o ma ion o bio ilms o dispe se hose al eady o med by pa hogens.
EPS p oduced by L. acidophilus ha e been demons a ed o inhibi he bio ilm o ma-
ion o a numbe o pa hogens, including en e ohemo hagic E. coli and S. en e i idis [
58
].
P e ious s udies ha e demons a ed ha he EPS om L. plan a um YW32 and L. aci-
Foods 2022,11, 1284 7 o 34
dophilus A4 we e ound o possess an i-bio ilm ac i i y agains bo h G am-nega i e and
G am-posi i e pa hogens [
59
,
60
]. In addi ion, HePS om L. e men um LB-69 [
61
] and L.
gasse i FR4 [
35
] displayed hei highes bio ilm inhibi ion on B. ce eus RSKK 863 and L.
monocy ogenes espec i ely.
An i-bio ilm ac i i y was also epo ed o o he LAB. Dex an p oduced by W. con usa
has been shown o ha e an ibio ilm ac i i y agains Candida albicans SC5314 [
62
]. L. ci eum
isola ed om bee sausages p oduced dex an and le an. These polysaccha ides showed
high ac i i y in dis up ing p e- o med bio ilms and bio ilm inhibi ion [63].
The quo um sensing (QS) sys em consis s o a mechanism in which bac e ia p oduce
molecules (gene ally oligopep ides in G am-posi i e bac e ia and acyl-homose ine lac one
in G am-nega i e bac e ia) by means o which hey de ec he size o densi y o he o he
bac e ia ha su ound hem and hus egula e he o ma ion o he bio ilm [
29
]. I has been
sugges ed ha EPS can ei he modi y he bac e ial coa and he eby hinde he a achmen o
bac e ia o su aces, o ac as a signaling molecule, and egula e gene exp ession in ol ed
in bio ilm o ma ion [
64
]. In his sense, as men ioned in he p e ious sec ion, se e al
sul a ed EPS exhibi ed a s onge inhibi o y e ec o mul iple G am-posi i e and nega i e
pa hogens han hose ee o sul a e. A possible eason o his is he in e up ion o he
signals ha media e bio ilm o ma ion o he e lux pa hway o wa e -soluble p o eins wi h
damage o he cell memb ane [29].
3.3. P ebio ic P ope ies
The e m p ebio ic was ecen ly de ined as a “subs a e ha is selec i ely u ilized
by hos mic oo ganisms con e ing a heal h bene i ” [
65
]. O ally deli e ed p ebio ics a e
“non-diges ible ood ing edien s h ough he gas ic and small in es ine, and when hey
each he la ge in es ine in humans, hey a e selec i ely u ilized by some in es inal bac e ia
popula ions, s imula ing he g ow h and/o ac i i y o in es inal mic obio a and he eby
wi h bene icial e ec on hos heal h” [
66
]. They a e gene ally poly- and oligosaccha ides,
such as inulin (
β
-(2
→
1)- uc an, galac o-oligosaccha ides and uc o-oligosaccha ides. The
me abolism o hese ca bohyd a es gene a es p oduc s, such as sho -chain a y acids,
gases, and o ganic acids, wi h posi i e e ec s on he hos , including o: (i) p o ide ene gy
o in es inal colonocy es, (ii) inhibi pa hogenic bac e ia, and (iii) modula e he human o
animal me abolism [6].
EPS syn hesized by LAB ha e a po en ial ole as p ebio ics as hey a e esis an o
GIT diges ion and can be selec i ely me abolized by bene icial gu bac e ia, especially
Bi idobac e ium spp. and Lac obacillus spp. [
67
]. The majo i y o LAB-EPS wi h demons a ed
p ebio ic po en ial a e HoPS and hei associa ed oligosaccha ides. I is possible ha he
mo e complex composi ion o HePS is esponsible o he low capabili y o he gu mic o-
bio a o hyd olyze and u he me abolize his ype o polyme s, limi ing hei p ebio ic
po en ial. Suppo ing his hypo hesis, i has been demons a ed ha he simple p ima y
s uc u e o HoPS allows ecal mic obio a o e men hem [
68
]. Dex an om W. ciba ia
RBA12 showed high esis ance o hyd olysis by a i icial gas ic juice,
α
-amylase and in es i-
nal luid and enhanced he g ow h o p obio ic Bi idobac e ia spp. and Lac obacillus spp. [
69
].
Le an om F. san anciscensis had a bi idogenic e ec and modi ied he gu mic obio a
composi ion [
70
]. In addi ion, a pu i ied
β
-glucan syn hesized by P. pa ulus 2.6 R, isola ed
om opy cide [
71
,
72
], imp o ed he g ow h o h ee p obio ic s ains L. plan a um WCFS1,
L. acidophilus NCFM, and L. plan a um WCFS1
β
-gal, which o e -exp ess a
β
-glycosidase
enzyme [
73
]. The p ebio ic e ec o a linea dex an p oduced by an L. pseudomesen e oides
s ain has ecen ly been in es iga ed by eeding mice wi h he polyme [
74
]. The esul s
e ealed ha he HoPS a ec ed he s uc u e o he gu mic obio a o he ea ed mice, by
dec easing he a io o Bac e oide es o Fi micu es [74].
Fu he mo e, some HePS om LAB which a e mainly composed o glucose, mannose,
galac ose and hamnose, ha e also shown p ebio ic ac i i y. A mannose- ich HePS p o-
duced by L. hamnosus GD-11 also s imula ed he g ow h o bi idobac e ia. Bo h HePS
ac ions ( -EPS1 and -EPS2) p oduced by L. delb ueckii subsp. bulga icus SRFM-1 p omo ed
Foods 2022,11, 1284 8 o 34
bi idobac e ia me abolism in a human ecal sample, gene a ing a high concen a ion o
sho -chain a y acids. Bo h EPS- ac ions we e mo e e ec i e han inulin in inc easing
popula ions o bi idobac e ia, lac obacilli and lac ococci [75].
3.4. Immunomodula o y E ec s
Commensal BAL ha e se e al bene icial oles in he GIT including con ibu ing o
main aining he in eg i y o he mucosal ba ie and p o ec ing agains pa hogens, as
well as p o iding nu ien s o euka yo ic cells. Thus, EPS molecules syn hesized by LAB
can adhe e o in es inal epi helial cells, he eby impeding pa hogen adhesion and/o
s imula ing he unde lying immune sys em cells [
76
]. The in e ac ion o he heal hy
mic obio a and he mucosal immune sys em is c ucial o he co ec de elopmen and
unc ion o he immune sys em. This p ocess akes place ia he pa e n ecogni ion
ecep o s (PRR) o hos cells, which in e ac wi h he molecula e ec o s ha a e p oduced
by in es inal mic oo ganisms. The immune sys em di e en ially ecognizes e ec o s
syn hesized by pa hogens and commensal bac e ia, including EPS, al hough hei s uc u es
a e simila , and hey sha e mechanisms o in e ac ion wi h he hos cells; his phenomenon
is called immune ole ance [
77
]. Thus, EPS p oduced by commensal and p obio ic bac e ia
(including LAB) a e in ol ed in he modula ion o (i) he inna e immune esponse by hei
in e ac ion wi h dend i ic cells and mac ophages, and (ii) he adap i e immune esponse,
by s imula ing he p oli e a ion o T and na u al kille cells. In bo h cases he modula ion
in ol es cy okine p oduc ion by he immune sys em o he hos euka yo ic cells, p o iding
di ec heal h-p omo ing bene i s, such as igh ing agains pa hogens and p e en ing o
GIT cance , as well as immunode iciency-induced diseases, such as in lamma o y bowel
diseases [25,78].
A numbe o published epo s on he
in i o
and
in i o
immunomodula o y o
immunos imula o y e ec s o HoPS and HePS om LAB ha e ecen ly been e iewed and
some o hese a e de ailed below [7,14,29].
In i o
expe imen s, including a compa a i e s udy o EPS-p oducing and non-
p oducing s ains, showed ha he high molecula mass HePS o he p obio ic L. casei
s ain Shi o a [
79
], L. hamnosus RW-9595M [
80
] and L. pa aplan a um BGCG11 [
81
] ha e
a supp essi e e ec on he ac i a ion o human mac ophages as a esul o inc easing
IL-10 and inhibi ion o no inc easing TNF-
α
, IL-1, IL-6 and IL-12. In addi ion, HePS and
sulpha ed-HePS, modi ied om S. he mophilus ASCC 1275, ha e an immunosupp essi e
esponse. Mo eo e , ea men wi h he HePS ASCC 1275 and sulpha ed-HePS ASCC 1275
esul ed in a dec ease in he p o-/an i-in lamma o y cy okines (IL-1
β
/IL-10, IL-6/IL-10,
and TNF-
α
/IL-10) sec e ion a ios o mu ine RAW 264.7 mac ophages s imula ed wi h
lipopolysaccha ide [
82
]. Simila ly,
in i o
assays demons a ed ha HePS om L. pa a-
plan a um CG11 had immunosupp essi e ac i i y in a mouse pe i oni is model induced by
ca ageenan, since he le els o he p o-in lamma o y media o s IL-1
β
, TNF-
α
and iNOS
dec eased, and inc eased sec e ion o he an i-in lamma o y IL-10 and IL-6 cy okines ook
place [83].
HePS ha e also been shown o exe immunos imula o y e ec s. Shin e al., [
84
]
ound ha HePS om P. pen osaceus KFT18 s imula ed NO and TNF-
α
, IL-6 and IL-1
β
p oduc ion in RAW 264
·
7 mac ophages and inc eased he p oli e a ion and he p oduc ion
o IL-2 and IFN-
γ
p ima y splenocy es
in i o
. In he same s udy, PE-EPS ea men
esul ed in an inc ease in he hymus and spleen lymphocy e and neu ophil coun in a
cyclophosphamide-induced immunosupp essed mouse model.
In i o
s udies in mice ha e e ealed ha o al adminis a ion o he HePS-p oducing
L. mesen e oides NTM048 s imula ed Peye ’s pa ch cells o induce IgA p oduc ion a he
in es inal and sys emic le els [
27
]. The au ho s epo ed ha EPS om NTM048 was an
immunos imulan ha enhanced mucosal IgA p oduc ion [
27
]. In addi ion, he in anasal
adminis a ion o EPS-NTM048 o mice wi h an an igen (o albumin) esul ed in he sec e-
ion o an igen-speci ic IgA and IgG in he ai way mucosa and he se um, sugges ing ha
he EPS has adju an ac i i y o use wi h mucosal accina ion [85].
Foods 2022,11, 1284 9 o 34
Wi h espec o HoPS, some s udies ha e demons a ed he abili y o hese polyme s
o modula e he immune esponse. Sa o e al. [
86
] ound ha an
α
-glucan (dex an) om L.
mesen e oides s ain exe ed immunos imula o y ac i i y. The le els o he IFN-
Foods 2022, 11, x FOR PEER REVIEW 9 o 35
p oduc ion in RAW 264·7 mac ophages and inc eased he p oli e a ion and he
p oduc ion o IL-2 and IFN-γ p ima y splenocy es in i o. In he same s udy, PE-EPS
ea men esul ed in an inc ease in he hymus and spleen lymphocy e and neu ophil
coun in a cyclophosphamide-induced immunosupp essed mouse model.
In i o s udies in mice ha e e ealed ha o al adminis a ion o he HePS-p oducing
L. mesen e oides NTM048 s imula ed Peye ’s pa ch cells o induce IgA p oduc ion a he
in es inal and sys emic le els [27]. The au ho s epo ed ha EPS om NTM048 was an
immunos imulan ha enhanced mucosal IgA p oduc ion [27]. In addi ion, he in anasal
adminis a ion o EPS-NTM048 o mice wi h an an igen (o albumin) esul ed in he
sec e ion o an igen-speci ic IgA and IgG in he ai way mucosa and he se um, sugges ing
ha he EPS has adju an ac i i y o use wi h mucosal accina ion [85].
Wi h espec o HoPS, some s udies ha e demons a ed he abili y o hese polyme s
o modula e he immune esponse. Sa o e al. [86] ound ha an α-glucan (dex an) om L.
mesen e oides s ain exe ed immunos imula o y ac i i y. The le els o he IFN- ϒ and IL-10
mRNA on mu ine splenocy es we e inc eased by he s imula ion. I was also demons a ed
ha dex an om L. sakei MN1 can up egula e he exp ession o IFN-1 and IFN-γ in ou
head kidney cells [87].
Fu he mo e, HoPS ha e also shown an i-in lamma o y ac i i y. Coloniza ion o he
gu o Lac obacillus- ee mice by he uc an-p oducing L. eu e i s ain 100-23, bu no by
he non-HoPS p oduce mu an , esul ed in inc eased p opo ions o egula o y T-cells
ma ked by exp ession o he ansc ip ion ac o Foxp3, and supp essed p oin lamma o y
T-cell esponses in he spleen [88]. In he same way, β-glucan om P. pa ulus 2.6 was able
o ac i a e human mac ophages wi h an an i-in lamma o y esponse [89]. In addi ion, he
exposu e o gno obio ic zeb a ish la ae o his HoPS caused he inhibi ion o gene
exp ession o he p o-in lamma o y cy okines, TNF-α and IL-8. Fu he mo e, he p o ein
adap o MyD88, which media es he ac i a ion o p o-in lamma o y cy okines ia NF-kB,
was inhibi ed [90]. Fu he mo e, he ec ui men and p oli e a ion o he neu ophils was
ep essed [88]. This β-glucan had an an i-in lamma o y e ec , which included a educ ion
in IL-8, bo h a he le el o i s gene exp ession and i s sec e ion, in an ex i o model o
human biopsies om pa ien s wi h C ohn’s disease [91].
The immunomodula o y ac i i ies o LAB-EPS a e con olled by hei
physicochemical p ope ies, such as monosaccha ide composi ion, molecula weigh ,
wa e solubili y, elec ic cha ges and s e eochemis y ( his opic will no be deal wi h he e
since i has been ecen ly e iewed in de ail by Xu e al. [7] and Zhou e al. [29]). Cu en ly,
he gene al opinion is ha he size and cha ge o polysaccha ides a e majo ac o s
in luencing he immune e ec . I seems ha nega i ely cha ged EPS and/o small size
molecules ac as s imula o s o immune cells, while neu al and la ge EPS ha e a
supp esso e ec . Mo eo e , many con adic ions exis wi h espec o his opic in he
e iewed li e a u e. This could be due o bo h he lack o de ailed knowledge o all EPS
s uc u es, as well as o he use o di e en in i o and in i o models o he
cha ac e iza ion o he immunomodula o y ac i i y o hese polyme s. In addi ion, mo e
s udies associa ing ac o s, such as monosaccha ide composi ion, unc ional g oups,
linkage pa e ns, and mic os uc u es o EPS wi h hei immune e ec , will be necessa y
o ully unde s and he s uc u e-immuni y ela ionship o LAB-EPS [7,29].
3.5. An i i al E ec s
I has been p oposed ha he an i i al ac i i y exe ed by p obio ics agains di e se
human and animal i uses may be media ed by mechanisms including he p oduc ion o
inhibi o y an i i al compounds, s imula ion o he immune sys em, and/o di ec
in e ac ion wi h i uses [92,93]. In pa icula , a ious EPS om p obio ic LAB could ha e
an i i al e ec s. Based on he mechanisms p oposed, hese e ec s could be conside ed as
(i) local o di ec , whe e he EPS may p e en i al in ec ion by blockage o i al
adso p ion by in e ac ion wi h ei he he i us pa icles o he hos cell [94], o (ii) sys emic
and IL-10
mRNA on mu ine splenocy es we e inc eased by he s imula ion. I was also demons a ed
ha dex an om L. sakei MN1 can up egula e he exp ession o IFN-1 and IFN-
γ
in ou
head kidney cells [87].
Fu he mo e, HoPS ha e also shown an i-in lamma o y ac i i y. Coloniza ion o he
gu o Lac obacillus- ee mice by he uc an-p oducing L. eu e i s ain 100-23, bu no by
he non-HoPS p oduce mu an , esul ed in inc eased p opo ions o egula o y T-cells
ma ked by exp ession o he ansc ip ion ac o Foxp3, and supp essed p oin lamma o y
T-cell esponses in he spleen [
88
]. In he same way,
β
-glucan om P. pa ulus 2.6 was
able o ac i a e human mac ophages wi h an an i-in lamma o y esponse [
89
]. In addi ion,
he exposu e o gno obio ic zeb a ish la ae o his HoPS caused he inhibi ion o gene
exp ession o he p o-in lamma o y cy okines, TNF-
α
and IL-8. Fu he mo e, he p o ein
adap o MyD88, which media es he ac i a ion o p o-in lamma o y cy okines ia NF-kB,
was inhibi ed [
90
]. Fu he mo e, he ec ui men and p oli e a ion o he neu ophils was
ep essed [
88
]. This
β
-glucan had an an i-in lamma o y e ec , which included a educ ion
in IL-8, bo h a he le el o i s gene exp ession and i s sec e ion, in an ex i o model o
human biopsies om pa ien s wi h C ohn’s disease [91].
The immunomodula o y ac i i ies o LAB-EPS a e con olled by hei physicochemical
p ope ies, such as monosaccha ide composi ion, molecula weigh , wa e solubili y, elec ic
cha ges and s e eochemis y ( his opic will no be deal wi h he e since i has been ecen ly
e iewed in de ail by Xu e al. [
7
] and Zhou e al. [
29
]). Cu en ly, he gene al opinion
is ha he size and cha ge o polysaccha ides a e majo ac o s in luencing he immune
e ec . I seems ha nega i ely cha ged EPS and/o small size molecules ac as s imula o s
o immune cells, while neu al and la ge EPS ha e a supp esso e ec . Mo eo e , many
con adic ions exis wi h espec o his opic in he e iewed li e a u e. This could be due
o bo h he lack o de ailed knowledge o all EPS s uc u es, as well as o he use o di e en
in i o
and
in i o
models o he cha ac e iza ion o he immunomodula o y ac i i y o
hese polyme s. In addi ion, mo e s udies associa ing ac o s, such as monosaccha ide
composi ion, unc ional g oups, linkage pa e ns, and mic os uc u es o EPS wi h hei
immune e ec , will be necessa y o ully unde s and he s uc u e-immuni y ela ionship
o LAB-EPS [7,29].
3.5. An i i al E ec s
I has been p oposed ha he an i i al ac i i y exe ed by p obio ics agains di e se
human and animal i uses may be media ed by mechanisms including he p oduc ion
o inhibi o y an i i al compounds, s imula ion o he immune sys em, and/o di ec
in e ac ion wi h i uses [92,93]. In pa icula , a ious EPS om p obio ic LAB could ha e
an i i al e ec s. Based on he mechanisms p oposed, hese e ec s could be conside ed as
(i) local o di ec , whe e he EPS may p e en i al in ec ion by blockage o i al adso p ion
by in e ac ion wi h ei he he i us pa icles o he hos cell [
94
], o (ii) sys emic o indi ec ,
as hese polyme s may indi ec ly hinde he i us by s imula ing he inna e and adap i e
immuni y o he hos cell [95].
This sec ion will discuss he s udies whe e i has been demons a ed ha he an i i al
e ec o EPS is o ally, o a leas pa ially, due o he ac ion o he immune sys em.
I has been shown ha EPS p oduced by LAB induce bene icial modula ion o he
sys emic and mucosal an i i al esponses and, in consequence, con ibu e o educing he
se e i y o i al in ec ions.
In i o
o al adminis a ion o yogu e men ed wi h he HePS-p oducing L. delb ueckii
OLL1073R-1 and he pu i ied HePS esul ed in a signi ican educ ion in in luenza i us
i e and a la ge inc ease in an i-in luenza i us an ibodies (IgA, IgG1). Fu he mo e, in
bo h g oups o ea ed mice, he ac i i y o na u al kille (NK) om splenocy es inc eased
signi ican ly [
96
]. In po cine in es inal epi helial cells (IECs), he inna e immune esponse,
Foods 2022,11, 1284 16 o 34
Table 1. Examples o use o LAB and hei applica ions on animals.
Food-P oducing Animals P obio ic LAB P obio ic Role Re e ences
Bo ine P oduc ion
Dai y Cows Lac ococcal (+p ebio ic)
P e en ing mas i is by diminishing in lamma ion o
mamma y gland and educing he le els o
mas i is-causing pa hogens (en e ococci and
s ep ococci).
[159]
Cal es
No mosil: L. b e is B-3,
L. plan a um 8 TIMES, L.
acidophilus 457,
En e ococcus aecium
UDS 86,
Inc easing he le els o no mal mic obio a, such as LAB
and bi idobac e ia, and dec easing he p e alence o
Esche ichia coli. Inc easing ed blood cells, he
concen a ion o hemoglobin, and γ-globulins wi hin
he physiological ange. Imp o ing phagocy ic eac ion
in he blood se um. These e ec s sugges a high
esponse o cal es o in ec ious agen s.
[160]
Cal es Lac obacillus u a um
LUHSS245
Inc easing and dec easing, espec i ely, le els o LAB
and en e obac e ia in eces. Posi i e in luence on ce ain
heal h pa ame e s in blood; educ ion in lac a e and
se um alanine amino ans e ase (AST) concen a ions.
[161]
Cal es
L.s casei DSPV 318T,
Lac obacillus sali a ius
DSPV 315T and
Pediococcus acidilac ici
DSPV 006T
Ea lie consump ion o s a e and, indi ec ly, p esumed
s imula ion o ea lie de elopmen o he umen,
omasum and e iculum, hus a o ing ea ly weaning.
Inocula ed cal es had be e g ow h pe o mance. This
beha io could be due o be e diges ion o lac ose and
sp ay-d ied whey p o eins.
[162]
Cal es
L. casei DSPV 318T, L.
sali a ius DSPV 315T
and P. acidilac ici DSPV
006T, L. plan a um
DSPV 354T.
The inocula we e added o a compu e ized milk eede
sys em. The e men a ion inc eased he shel li e o he
milk, a oiding, in consequence, he need o equen ly
disca d he milk wi h a cos educ ion. The inocula
s imula ed milk in ake. Lac obacillus/coli o m a io was
>1 in he p obio ic g oups and <1 in he con ol g oup.
De ec ion o inc eased gain o body weigh gain and
eed e iciency.
[163]
Swine P oduc ion
Sows
and Pigle s
L. eu e i,L. amylo o us,
and L. johnsonii
Sows
Imp o emen in ep oduc i e pe o mance including
inc eased numbe o bi h pigle s and bi h weigh pe
li e , concep ion a e du ing es us, and lowe numbe s
o weak pigle s. Inc eased an ioxidan
capaci y—concen a ion o malondialdehyde ac i i y
(MDA) was lowe in he g oup ed wi h he LAB.
Inc easing immune indexes—TNF-and IgA we e highe
compa ed wi h he con ol.
Pigle s
Imp o emen o g ow h de elopmen and dec ease in
dia hea incidence—LAB inc eased inal body weigh
and dec eased incidence o dia hea. Inc ease in a e age
daily gain, daily eed in ake, and he e iciency o eed
u iliza ion o pigle s in he g oup ed wi h he LAB.
Inc eased an ioxidan ac i i y—concen a ions o
supe oxide dismu ase (T-SOD) we e signi ican ly highe
in he LAB- ea ed g oup and he MDA concen a ion
was lowe . Inc eased immune indexes—TNF, IgA, IgG
we e highe in he g oup ed wi h LAB han he con ol.
[164]
Pigle s L. sali a ius The inocula inc eased he numbe o lac obacilli and
illus heigh in he duodenum, jejunum, and ileum. [165]
Foods 2022,11, 1284 17 o 34
Table 1. Con .
Food-P oducing Animals P obio ic LAB P obio ic Role Re e ences
Poul y
Poul y
En e ococcus aecium LET
301, L. sali a ius LET 201,
L. eu e i LET 210,
P e en ed nega i e e ec s o an inu i ional ac o s,
such as die a y lec ins soybean agglu inin (SBA) and
whea ge m agglu inin (WGA). The ea men wi h he
bac e ia p o oked an inc ease in se e al diges i e
enzyme ac i i ies and alkaline phospha ase (an
in es inal ma u a ion ma ke ) in bi ds ed wi h a
con en ional die . In addi ion, supplemen a ion wi h
he bac e ia coun e ac ed: (i) he dele e ious e ec s o
SBA inc eased con en o SBA, and (ii) he nega i e
e ec o a WGA die a y sou ce on he ac i i y o
diges i e enzymes and in es inal mucosa in eg i y.
[166]
Eggs L. lac is subsp. c emo is
In he spleen and cecal onsils o b oile chickens,
p ebio ics o synbio ics p oduced by he BAL s imula ed
gene exp ession in ol ed in ene gy me abolism and
immune esponse.
[167]
B oile L. sali a ius and
P. pa ulus
Imp o emen in immune esponse, bone cha ac e is ics,
weigh gain and in es inal mo phology, as well as
dec ease in Salmonella en e i idis coloniza ion.
[168]
B oile s (eggs)
Bi idobac e ium bi idum,
G3; B. animalis,G4; B.
longum,G5; o B.
in an is,G6
T ea men wi h bi idobac e ial, ins ead o LAB,
inc eased b oile g ow h pe o mance de ec ed by body
weigh , weigh gain and a io o eed con e sion.
Enhanced hy oid ho mone me abolism since
concen a ions in se um o hy oxin and
iiodo hy onine we e ele a ed upon bac e ial ea men .
Ileal a chi ec u e was imp o ed: highe illus heigh
alues and he illus heigh /c yp dep h a io. The
le els o ileal LAB and Bi idobac e ium spp. inc eased. By
con as , o al coli o m le els, as well as bac e ial
coun s, dec eased.
[169]
B oile s Lac obacillusBi idobac e ia
Imp o ed body weigh gain and p e en ion o he
dele e ious and/o le hal e ec s o Salmonella in ec ion
in chicks by wo mechanisms: compe i i e exclusion
and he enhancemen o cy okine sec e ion.
[170]
B oile s L. sali a ius DSPV 001P Imp o ed body weigh and endency o educe he a e
o mo ali y. [171]
Sheep p oduc ion
Sheep L. hamnosus Rumen mic obiome s uc u e and abundance we e
sligh ly al e ed. [172]
Goa p oduc ion
Goa L. hamnosus and
E. aecalis
Enhanced weigh gain and d op in he gu pH, hus
main aining an equilib ium o uminal mic obio a [173]
Aquacul u e
Ju enile con ic
cichlid ish L. casei PB-LC39
Imp o ed g ow h: inal body weigh , pe cen age o
weigh gain (%), speci ic g ow h a e, ood con e sion
a io, and p o ein con en o whole-body composi ion
we e signi ican ly highe . Inc eased ac i i y o diges i e
enzymes (p o ease, amylase, and lipase). Imp o ed
immune esponse (le els o o al immunoglobulin (Ig),
and se um globulin we e inc eased). Changes in he
mic obio a: le els o LAB in ish gu we e enhanced.
A e an ai -di e es , he a es o ish eco e y in he
g oup ed wi h he LAB was signi ican ly highe han
ha in he con ol g oup.
[174]
Foods 2022,11, 1284 18 o 34
Table 1. Con .
Food-P oducing Animals P obio ic LAB P obio ic Role Re e ences
Snakehead ish
(Channa a gus)
L. lac is L19 and E.
aecalis W24
Inc eased eed e iciency a io, speci ic g ow h a e, inal
body weigh , weigh gain, and p o ein e iciency a io.
Inc eased IgM, ACP, AKP, LZM, C3 and C4 ac i i y in
se um, which could induce humo al immuni y.
Up- egula ion o he exp ession o IL-1β, IL-6, IL-10,
TNF-α, IFN-γ, HSP70, HSP90, TGF-βin he spleen,
head kidney, gill, li e and in es ine. A e challenge
wi h Ae omonas e onii, inc eased su i al a es and
esis ance o disease.
[175]
Apicul u e
Honey bee L. b e is B50 Biocenol
Inc eased esis ance o in ec ious diseases and s ess
condi ions: inc ease in he a io o lac ic acid bac e ia o
en e obac e ia. Enhanced immuni y in bee colonies:
inc eased exp ession o an imic obial pep ides (abaecin,
de ensin-1) coding genes and pa e n ecogni ion
ecep o s (Toll-like ecep o and pep idoglycan
ecogni ion p o eins).
[176]
Howe e , some o hese epo ed p obio ic lac obacilli s ains, L. eu e i,L. amylo o us,
and L. johnsonii, we e selec ed, among o he p obio ic cha ac e is ics, o hei high capaci y
o p oduce EPS, and hey we e used o analyze hei e ec on eeding o sows and weaned
pigle s. The esul s ob ained indica ed ha hese LAB con ibu ed o imp o emen in he
ep oduc i e pe o mance o sows and weaned pigle g ow h, p esumably due in bo h o a
posi i e e ec on an ioxidan enzyme ac i i y and in he immune indexes (Table 1) [
164
].
Al hough he au ho s did no di ec ly co ela e he biological ac i i ies o he p oduc ion
o EPS, i could be due o he syn hesis o hese polyme s, as discussed in he p e ious
sec ion, and as ecen ly epo ed o he EPS om L. hamnosus GG, which has been
shown o be an e ec i e d ug o elie e oxida i e s ess [
177
]. On he o he hand,
in i o
assays demons a ed ha he lac obacilli men ioned abo e showed good inhibi ion agains
en e o oxigenic Esche ichia coli (ETEC), which exp essed he imb iae K88. Simila ly, o he
au ho s ha e epo ed ha L. eu e i s ains p oduce eu e an o le an, EPS ha inhibi he
ETEC binding o he mucosa, and ca ied ou an
in i o
s udy o eed e men a ion. They
concluded ha he L. eu e i eed e men a ion dec eased he coloniza ion o weaning pigle s
by he ETEC and ha he eu e an p o ided by he LAB may con ibu e o p e en ing
ETEC adhesion o he in es inal mucosa [
178
]. La e , an
in i o
s udy demons a ed ha
glucan polyme s om L. eu e i we e in ol ed in supp ession o he in lamma o y esponse
o ETEC in ec ion. This s udy ea ed po cine epi helial cells wi h he glucan p oduced
by L. eu e i, and he global esponse a he le el o egula ion o gene exp ession was
analyzed by high- h oughpu RNA-sequencing [
179
]. Mo eo e , al hough u he
in i o
s udies a e needed,
in i o
an i i al ac i i y was demons a ed o L. delb ueckii TUA4408L
and i s HePS, which was able o enhance he esis ance o po cine in es inal epi helial
cells o o a i us in ec ion by modula ion o he inna e an i i al esponse and educ ion in
i al eplica ion [
97
]. Recen ly, EPS om L. plan a um p e en ed he adso p ion o po cine
epidemic dia hea i us, and diminished inju ed Ve o cells induced ea ly apop osis, as well
as in lamma o y esponses [180].
Ke i g ains a e a ese oi o p obio ic LAB, and hey ha e been used expe imen ally
in slaugh e animals. In he mic obio a o he ke i g ains, he mos ep esen a i e species
a e Lac obacillus ke i ano aciens,Len ilac obacillus ke i i and Len ilac obacillus pa ake i i. How-
e e , o he LAB a e also p esen (L. pa acasei,Lac obacillus acidophilus,L. delb ueckii subsp.
bulga icus,L. plan a um and L. lac is) [
181
,
182
]. Thus, Bengoa e al. [
181
] ha e ecen ly
e iewed he in luence o he EPS o L. pa acasei on he p obio ic p ope ies o his BAL.
Due o he ins abili y o he mic oo ganisms ha make up he g ain, hei cha ac e iza ion
Foods 2022,11, 1284 19 o 34
o e ime showed a iable esul s. Bo h ex insic and in insic ac o s ha a ec mic obial
g ow h ha e a di ec in luence on he mic oo ganisms ha make up he g ain, which is
why hey a e usually classi ied as compe i i e exclusion cul u es (CEC), a concep p e i-
ously desc ibed by Nu mi and Ran ala [
183
]. The p inciple o his s a egy is based on he
compe i ion be ween he no mal mic obio a, he non-pa hogenic in es inal mic obio a and
he pa hogenic bac e ia o colonize he GIT o he hos [184]. CEC equi es he addi ion o
a non-pa hogenic bac e ial cul u e o he in es inal ac o slaugh e animals o dec ease
coloniza ion by popula ions o pa hogenic bac e ia [
185
]. CEC ha e been shown o be an
e ec i e me hod o he con ol o salmonellosis in comme cial poul y [
186
]. Depending
on he s age o p oduc ion (s a e o gu ma u i y), he objec i e o CEC may be he exclusion
o pa hogens om he gu o he newbo n animals, o he displacemen o an al eady
es ablished popula ion o pa hogenic bac e ia [
187
]. A no el app oach is o use his ool o
educe he in es inal bu dens o a bac e ium ha is a no mal hos inhabi an bu pa hogenic
o humans, such as he mo ole an Campylobac e in poul y. Ke i con ains Lac obacillus
spp. and yeas s and may be a use ul CEC candida e o imp o e he in es inal mic obio a o
poul y and he hygiene o he ca casses ob ained om hem [
188
]. The bene icial e ec can
be a ibu ed o he inhibi ion o pa hogenic mic oo ganisms by me aboli es (e.g., o ganic
acids sec e ed by LAB belonging o he ke i g ain mic obio a). I has also been epo ed
ha he S-laye p o ein o Lac obacillus ke i could ha e a p o ec i e ole agains Salmonella
en e i idis [
189
]. Some
in i o
s udies on he use o ke i in slaugh e animals ha e been
epo ed wi h encou aging esul s [
190
–
192
]. Howe e , he bene icial e ec indica o s,
ela ed o bo h g ow h pe o mance and animal heal h, should con inue o be s udied
o each mo e obus conclusions. The limi a ions and equi emen s o ials wi h big
animals ha mus be ed wi h esh p obio ic p oduc s in hei complex p oduc ion sys ems
may explain he lack o epo s in he li e a u e. Howe e , he pe o mance o such es s
should be s imula ed because he adminis a ion o ke i o young cal es could gene a e a
bene icial e ec on p oduc ion and heal h a es and on he immune sys em o animals.
The e a e o he examples o EPS-p oducing p obio ic LAB assayed in poul y. The L.
johnsonii FI9785 s ain was shown o be e ec i e as a compe i i e exclusion agen agains
Campylobac e jejuni. A educ ion in he coloniza ion o C. jejuni was obse ed and he
composi ion o he in es inal mic obio a was signi ican ly al e ed [
193
]. In a p e ious s udy,
he s uc u e and biosyn hesis o wo EPS (dex an and HePS) om his p obio ic s ain
we e epo ed, as well as he p obable ole o hese polyme s in bio ilm o ma ion, and hos
coloniza ion as p o ec ion agains he hos ile en i onmen o he in es ine [48,50].
The p obio ic s ain L. gasse i F4 isola ed om he GIT o ee- ange chickens p o-
duced a HePS. The an ioxidan , an ibac e ial and an ibio ilm ac i i ies, es ed
in i o
,
we e a ibu ed o ha polyme , so he esea che s sugges ed i s po en ial use as a ood
addi i e [35].
Ano he s udy demons a ed he p oduc ion o EPS in ou s ains o L. sali a ius
isola ed om chicken eces (HoPS and also HePs in one o hem). These s ains p oduced
di e en amoun s o EPS, which in u n a ec ed he
in i o
capaci ies o bio ilm o ma ion,
au oagg ega ion and adhesion. The au ho s highligh ed ha all s ains exhibi ed inhibi o y
e ec s agains chicken pa hogens [194].
Thus, he use o EPS-p oducing p obio ic LAB in he die o ood-p oducing animals
has g ea po en ial o p e en o educe he sp ead o pa hogens du ing he p ima y
p oduc ion s age and could be an e ec i e al e na i e ool o he use o an imic obials in
p ima y ood p oduc ion.
6. Applica ions o EPS o he Imp o emen o Technological P ope ies o Food
Fe men a ion wi h EPS-p oducing LAB is used pa icula ly in he ood indus y since
he in si u p oduc ion con ibu es o imp o ing he o ganolep ic quali y, senso y and
heological p ope ies, as well as he s abili y, o he inal p oduc s. Fu he mo e, he in
si u p oduc ion o EPS has he ad an age o educing he quan i y o s abilizing agen s
used o imp o e ex u al p ope ies in dai y and e men ed ce eal p oduc s. Recen ly,
Foods 2022,11, 1284 20 o 34
bo h in si u p oduc ion and inco po a ion in o oods has led o hem being conside ed as
unc ional ing edien s, due in pa o hei pos ula ed heal h bene i s, as de ailed abo e,
and unc ionali y in he ood i sel [12,13].
In gene al, EPS a e used in he ood indus y mainly as: (a) iscosi y agen s, (b) ex u e,
mou h eel, and eeze- haw s abili y enhance s, (c) hickene s and so ene s, (d) salad
d essings, and (e) ilms and coa ing agen s, o men ion a ew [
130
,
195
]. These applica ions o
EPS a e due o hei physical and heological p ope ies which emain s able unde di e en
indus ial p ocessing condi ions [
52
,
63
]. Fu he mo e, ce ain ecological cha ac e is ics,
such as hei biodeg adabili y and non- oxici y, ha e inc eased hei applica ions as na u al
emulsi ie s compa ed o chemically syn hesized p oduc s, which a e commonly associa ed
wi h a nega i e impac on he en i onmen [
196
]. Ne e heless, i has been sugges ed ha
many o hese p ope ies depend s ongly on he amoun o EPS p esen in ood p oduc s,
hei chemical s uc u e, unc ional g oups and na u e o glycosidic bonds which may
in e ac in di e en ways wi h he ood ma ix a ec ing he physical, heological and
ex u al cha ac e is ic o he inal p oduc [14,197,198].
6.1. Dai y P oduc s
In gene al, EPS ha e been employed by he dai y indus y o educe he pe cen age
o added milk solids, since hey can be used as hickene s and s abilize s ha imp o e
he s uc u e and consis ency o oods, wi hou modi ying o ganolep ic p ope ies [
199
].
Fu he mo e, EPS addi ion a oids he syne esis o e men ed milk p oduc s, such as yogu
and cheese, e en upon p oduc s o age. In he case o yoghu , he common s a e cul u es
Lac obacillus delb ueckii subsp. bulga icus and S. he mophilus ha e been selec ed due o hei
abili y o p oduce HePS in he ange o 60–150 mg/L and 30 mg/L, espec i ely [
200
,
201
].
Recen ly, new possible s a e s o adju an s ha e been s udied o be applied in yogu and
cheese manu ac u ing.
An al e na i e, o coun e ac he p oblems associa ed wi h low a con en in e men ed
dai y p oduc s, is he use o s a e s wi h high in si u EPS p oduc ion, as has been desc ibed
o he HePS p oduce Limosilac obacillus mucosae DPC 6426 ( o me ly Lac obacillus mucosae),
which imp o ed he unc ional and heological p ope ies du ing he p oduc ion o a
low- a yoghu , as well as signi ican ly dec easing syne esis [
135
]. In he same manne ,
low- a con en impac s on he ex u e and de elopmen o he la o o cheese [
14
,
202
],
and, as in he case o yogu , an EPS-p oduce added as s a e o adju an cul u e, has
been shown o imp o e he ex u e and quali y o a - educed cheeses [
200
]. In low- a
mozza ella, and semi- a o low- a chedda cheese, he addi ion o some HePS-p oducing
s ains o he genus Lac obacillus (L. delb ieckii subsp. bulga icus MR-1R), in combina ion wi h
S. he mophilus and Lac ococcus lac is ssp. c emo is DPC6532, inc eased mois u e con en and
yield, while imp o ing i s mel ing, bu had no nega i e impac on la o , when compa ed
wi h con ol cheeses [203,204].
A s udy o di e en EPS p oduced by di e en s ains o L. lac is, in a model o a
low- a esh cheese, e ealed ha , in di e en ways, hey modi ied he s uc u al and
mac omolecula p ope ies o he p oduc . In his model, one s ain, which p oduced a
opy capsula EPS (LL-1+), p o ided highe gel s i ness, inc eased wa e e en ion, lowe
pa icle size and imp o ed c eaminess ex u e, in compa ison o a non- opy EPS (LL-2)-
p oducing s ain [
205
]. Nguyen e al. [
19
] epo ed ha , o a oid some quali y de ec s in
yogu ex u e (g aininess), ce ain c i e ia mus be conside ed, such as he mo phology
o he bac e ia and he s uc u e o he EPS p oduced by hem, as well as he inal EPS
concen a ion and he a e o acidi ica ion in he p oduc .
In Scandina ian e men ed milk d inks, such as iili, ae e, il, and sky , he heological
cha ac e is ics, such as i mness, hickness and sliminess, ely on he abili y o opy s ains
o L. lac is subsp. lac is and L. lac is subsp. c emo is o p oduce di e en HePS ha con e
ex u e o he inal p oduc [206].
Fo he p oduc ion o ice-c eam a indus ial scale, indispensable addi i es o i s man-
u ac u e, such as gums s abilize s, galac omannan hyd ocolloids o chemically modi ied
Foods 2022,11, 1284 21 o 34
plan ca bohyd a es (s a ch, pec in, gua gum, e c.) and glucomannan based “salep”, a e
equi ed. Howe e , EPS ha e shown some unc ional p ope ies as s abilize s, and i may
be possible o eplace hem by in si u EPS p oduc ion by LAB. De li e al. [
207
] analyzed
he impac o in si u HePS p oduc ion by S. he mophilus s ains on di e en echnolog-
ical and senso ial p ope ies o ice c eam, in an e o o de elop a unc ional ice-c eam
wi hou adding common s abilize s. A heological analysis o he ice-c eam showed an
imp o emen in iscosi y which was associa ed wi h EPS p ope ies as a hickening and
gelling agen , while o he echniques demons a ed he p esence o a web-like compac
mic os uc u e wi h holes ha also co ela ed wi h he imp o ed heological p ope ies.
6.2. Mea P oduc s
Since ea ly imes, o ganic acids, such as lac ic acid and ace ic acid, ha e been employed
as he p incipal na u al me hod o p ese ing aw mea . Lac obacillus and Pediococcus a e
he main gene a used o imp o ed ood sa e y in aw mea , as hey p oduce lac ic and ace ic
acids which dec ease he popula ion o o he indigenous bac e ia. Bac e iocin o nu ien
compe i ion may also con ibu e o his sa e y, and he de elopmen o colo and ex u e,
as well as o he cha ac e is ics o mea p oduc s [
129
]. In he mea indus y, hyd ocolloids
and phospha es a e he main ing edien s in en ionally inco po a ed o enhance he quali y
o mea p oduc s [
208
]. Hyd ocolloids ha e been used o imp o e wa e holding capaci y,
emulsion s abili y, and in luence he gelling p ope ies o mea p o eins, yield and juiciness
o cooked p oduc s, sp ead abili y and mou h eel in a - educed p oduc s o o upg ade
he ex u al cha ac e is ics o low o educed a con en [119,209,210].
Nowadays, consume demand o low- a and/o addi i e- ee mea p oduc s is in-
c easing, so he de elopmen o new p oduc s in he mea indus y is impo an in his
ma ke a ea. Howe e , i can also cause d awbacks as low- a p ocessed mea p oduc s
ha e some de ec s ega ding hei echnological and senso y quali ies [
129
,
211
]. In his
con ex , in si u EPS p oduc ion by LAB seems a p omising al e na i e. Few s udies ha e
desc ibed analysis o his p oduc ion in mea p oduc s, including cooked ham, aw e -
men ed sausages, and sp eadable aw e men ed sausages, in which he polyme s enhance
wa e -binding capaci y o educe he amoun o a added in o he p oduc s. This las e ec
will be bene icial o human heal h, since inges ion o high- a mea p oduc s is associa ed
wi h diabe es and ca dio ascula disease [208].
De li e al. [
207
] e alua ed he e ec o wo EPS p oduce s (HoPS and HePS p oduce s,
espec i ely), Lac iplan ibacillus plan a um 162 R ( o me ly Lac obacillus plan a um) and
Leuconos oc mesen e oides N6 on a Tu kish e men ed sausage. The inal p oduc s showed an
imp o emen in hei ex u al p ope ies (less s icky, ha de and oughe wi h a web-like
s uc u e in he sausage ma ix) in compa ison wi h hose sausages con aining a non-EPS
p oduce . Simila ly, he in si u HePS p oduc ion by L. plan a um TMW 1.1478 was e iden
on he inal p ope ies o salami (a d ied ype o e men ed sausage), which con ibu ed
nega i ely o he quali y a ibu es, as i was signi ican ly so e despi e no been a ec ed
in e ms o i s senso ial p ope ies [212].
Addi ionally, in he esea ch by Hilbig e al. [
213
], he e ec o wo EPS was e alua ed
on he sp ead abili y o a a - educed e men ed aw sausage (“Teewu s ”). Lac ilac obacillus
sakei TMW 1.411 ( o me ly Lac obacillus sakei), as well as Lac ilac obacillus cu a us TMW
1.1928 ( o me ly Lac obacillus cu a us), bo h HoPS p oduce s, we e inocula ed as s a e
cul u es in a aw sausage model. The inal p oduc s e ealed ha EPS p oduced by
bo h s ains du ing e men a ion made i possible o educe a ound 20% o a con en ,
he eby signi ican ly imp o ing he so ness and sp ead abili y in hose sausages wi h
EPS compa ed o con ol samples con aining non-EPS p oduce o inocula ed wi h he
HePS p oduce , L. plan a um TMW 1.1478, which had a nega i e e ec on he quali y o
he p oduc .
Foods 2022,11, 1284 22 o 34
6.3. Fe men ed Be e ages
The o ganolep ic and senso ial cha ac e is ics o some e men ed be e ages, such as
ke i and pulque, ha e been a ibu ed o LAB and o hei EPS p oduc ion in si u du ing
he e men a ion p ocess [12,195].
Ke i , a sel -ca bona ed and sligh ly alcoholic dai y be e age consumed in Eas e n
Eu ope, is p epa ed om ke i g ains, which a e mainly composed o p o eins and polysac-
cha ides, as well as a combina ion o LAB (homo- and he e o- e men a i e), ace ic bac e ia
and yeas , a symbio ic conso ium cha ac e is ic o g ains [
182
,
214
]. Du ing e men a ion,
pep ides and polysaccha ides (ke i an) a e p oduced, ac ing as iscosi y agen s. Ke i an, is
a soluble glucogalac an composed o equi alen pa s o glucose and galac ose, which has
been epo ed o ha e a molecula mass be ween 10
5
and 10
7
Da [
132
,
215
], and is mainly
p oduced by Lac obacillus ke i ano aciens and o he lac obacilli [
216
]. O he EPS-p oducing
LAB ha e been isola ed om ke i g ains, such as L. plan a um,Lac icaseibacillus pa acasei
( o me ly Lac obacillus pa acasei), Lac obacillus hel e icus ( o me ly Lac obacillus sun o yeus),
Lac iplan ibacillus pen osus ( o me ly Lac obacillus pen osus), L. lac is subsp. lac is and L.
mesen e oides [217,218].
Pulque, is an alcoholic be e age made om he e men a ion o aguamiel, he esh
sap ex ac ed om di e se aga e species, and is consumed mainly in hose egions in
Mexico whe e hese species o aga e a e cul i a ed. The inal senso y cha ac e is ics in
pulque a e de e mined by he e men a ion ime o he aguamiel, he inc emen in iscosi y
as a consequence o he p oduc ion o EPS and he deg ee o alcohol p oduced [
219
]. L.
mesen e oides IBT-PQ, has been ecognized as one o he mos impo an mic oo ganisms
in pulque e men a ion, p oducing a soluble linea dex an wi h glucose molecules linked
by
α
-(1,6) bonds wi h b anching om
α
-(1,3) bonds syn hesized om suc ose p esen in
aguamiel and pulque. O he Leuconos oc species, such as Leuconos oc ci eum and Leuconos oc
kimchi, ha e also been epo ed as he mos abundan LAB species du ing he ea ly s ages o
pulque e men a ion, so i has been p oposed ha di e en EPS p oduc ion may con ibu e
o he o ganolep ic cha ac e is ics o bo h be e ages [219].
In e men ed soymilk p oduc s, EPS-p oducing LAB ac as unc ional s a e cul u es
as hey con ibu e o he consis ency and heological p ope ies [
220
]. Va ious soybean-
based p oduc s, such as soy sauce and soy pas e, ha e ecen ly been p oduced using
EPS-p oducing LAB [
198
]. Li e al. [
221
] moni o ed he e men a ion o e 21 days o soy
milk wi h wo EPS-p oducing L. plan a um 70810 and L. hamnosus 6005 s a e s. The
e men ed soy milk wi h L. plan a um 70810 conse ed he iscosi y p o ile, inc eased he
echnological p ope ies and enhanced he mou h- as e o soy milk.
6.4. Ce eal-Based Be e ages and Food P oduc s
Ce eal-associa ed LAB p oduce a wide a ie y o EPS and oligosaccha ides due o
glycansuc ase ac i i y, enhancing bo h he ex u al and o ganolep ic cha ac e is ics o he
inal p oduc . I is easible o use hem as p obio ics and hei polyme s as p ebio ics
and/o pos bio ics. In his con ex , Pé ez-Ramos e al. [
76
] e alua ed he abili y o p oduce
β
-glucan by he P. pa ulus 2.6 R ( opy s ain) and he isogenic non-p oducing s ain P.
pa ulus 2.6 NR (non- opy), in h ee di e en ce eal-based ma ices. The le els o
β
-glucan
we e highe in oa and ice lou s e men ed wi h he opy s ain and enhanced heological
p ope ies in he inal p oduc we e obse ed.
Lo usso e al. [
222
] e alua ed quinoa lou -based e men ed be e ages wi h di e -
en LAB, including he p obio ic L. hamnosus SP1, an EPS-p oducing Weissella con usa
DSM20194, and he L. plan a um TB610 s ain isola ed om quinoa. A e 20 h o e -
men a ion, inc emen in he iscosi y and wa e -holding capaci y o he quinoa be e age
due o a dex an- ype EPS p oduced by W. con usa was obse ed. In addi ion, a s able
EPS–p o ein ne wo k which also con ibu ed o enhanced ex u al p ope ies o he p oduc
was egis e ed.
Foods 2022,11, 1284 23 o 34
6.5. Dough and Bake y P oduc s
The wo main unc ions o hyd ocolloids inco po a ed in o dough sys ems and b eads
a e: (i) he wa e holding capaci y and wa e dis ibu ion, and (ii) s uc u al in e ac ions
wi h componen s such as glu en, non-glu en p o eins, and s a ch. Addi ionally, hyd ocol-
loids may s abilize he in e ace o he dough liquid ilm su ounding gas bubbles, hus
imp o ing gas e en ion [223].
HoPS p oduce s a e usually inco po a ed in p oduc s such as sou dough, as hey ha e
a ce ain amoun o suc ose, which se es as a ca bon sou ce o he in si u p oduc ion
o hese ype o EPS [
206
], and in luences he s uc u al quali y and baking abili y in he
bake y. Dex an p oduced by bac e ial e men a ion and added a a le el below 5% as a
ood ing edien in he baking indus y was app o ed by he Eu opean Commission in 2000,
on he basis ha i does no cons i u e a haza d o consume heal h [224].
The use o EPS p oduce s a e s has a ac ed he a en ion o he bake y indus y, as
an inc ease in demand o glu en- ee p oduc s mus be me . Howe e , baking wi hou
glu en, he main ing edien ha in luences he s uc u e and quali y o b ead, con inues
o be a challenge. In addi ion, ce eals ee o glu en a e employed o p oduce glu en- ee
(GF) p oduc s. Glu en- ee g ains include so ghum, ice, co n, mille and e , and he
pseudoce eals, ama an h, quinoa and buckwhea [
225
]. Howe e , he lou s ob ained om
hese g ains lead o p oduc s wi h poo s uc u e, ex u e and mou h eel. GF b eads ha e
low wa e abso p ion, app eciable changes in c umb ea u es, olume educ ion, and poo
s abili y o b ead. These de ec s could be p e en ed by inco po a ion o EPS-p oducing
LAB in o glu en- ee sou dough [6].
Some e idence sugges s ha EPS, such as dex an p oduced by L. mesen e oides and
W. ciba ia, le an om F uc ilac obacillus san anciscensis, ( o me ly Lac obacilllus san ancis-
censis), and eu e an om L. eu e i, clea ly a ec dough heology and b ead ex u e, and
i may be possible o eplace o educe he use o hyd ocolloids in whea and glu en- ee
baking [201,226].
The abo e was shown by Ka ina e al. [
227
] du ing he in si u dex an
p oduc ion by W. con usa VTT E-90392 in whea sou dough, which imp o ed he iscosi y
o he sou dough, and inc eased he inal olume o he b ead (a ound 10%) and he c umb
so ness (25–40%) wi hou p oducing s ong acidi y. Simila esul s we e epo ed o
he in si u syn hesis o dex an by W. ciba ia MG1 in glu en- ee sou doughs (buckwhea ,
quinoa, and e ) and a W. con usa on a whole g ain pea l mille b ead, in which he dex-
an p oduced imp o ed b ead olume, educed c umb i mness and a oided mois u e
los [228,229].
Fu he mo e, due o an inc eased demand o uncon en ional lou s (e.g., legumes,
pseudoce eal, e c.), sou doughs p epa ed wi h unusual g ains migh be a good sou ce o
EPS-p oducing s ains. Legume-base sou dough seems a po en ial al e na i e, despi e he
lack o glu en in lou s ob ained om hese, which can be o e come by he in si u p oduc ion
o EPS du ing he e men a ion p ocess [
230
]. W. con usa Ck15, a dex an p oduce , was
isola ed as a dominan s ain a e se e al backsloppings o a spon aneously e men ed
chickpea lou dough. In si u dex an p oduc ion by W. con usa Ck15 e men a ion imp o ed
he iscosi y and he p oduc ion o he EPS pe cen age in he doughs wi h espec o he
o he doughs inocula ed wi h ano he dex an p oduce , Leuconos oc pseudomesen e oides
DSM 201193, and a non-EPS p oduce L. plan a um F8 [230].
7. Fu u e Pe spec i es
The use o EPS-p oducing LAB and hei polyme s in he dai y indus y is well
es ablished, and hei usage in he de elopmen o mea and ce eal-based ood is p edic ed
o expand in he nea u u e. These oods will be used p incipally o g oups o he
popula ion ha equi e low- a o glu en- ee e men ed p oduc s.
The sea ch o new EPS p oducing LAB will be acili a ed by he ac ha many o
hem can be isola ed om he GIT and/o animal eces, and he use o hese bac e ia will
place emphasis on hei applica ion acco ding o hei o igin. Ne e heless, conce ning he
isola ion and pu i ica ion o LAB-EPS, special ca e should be aken o p e en deg ada ion
Foods 2022,11, 1284 24 o 34
o he polyme s du ing hei ex ac ion. Special measu es should also be aken o a oid
he p esence o any impu i ies, such as lipopolysaccha ides, which can in e e e wi h he
cha ac e iza ion o he polyme ’s biological ac i i y. Fu he mo e, EPS s uc u es can
di e signi ican ly be ween bac e ial s ains and esul in di e en biological p ope ies.
De ails o he speci ic EPS s uc u es used in esea ch a e o en lacking o a e insu icien
o de e mine he exac s uc u e– unc ion ela ionships. When epo ing indings on he
biological unc ions o EPS, special ca e should be aken in p o iding de ailed in o ma ion
on he bac e ial s ain used, and he speci ic EPS s uc u e.
The li es ock indus y is impo an wo ldwide, and he a ming indus y is demanding
imp o emen o animal eed wi h p ebio ics and pos bio ics. Ne e heless, he ma ke
o eeds u LAB is s ill in an ea ly s age o de elopmen and cu en ly he numbe and
olume o comme cialized p oduc s con aining LAB is ela i ely small. Mo eo e , he e
a e no p obio ic EPS-p oducing LAB no bac e ial EPS cu en ly used in animal heal h.
Thus, his is a new ield which dese es o be explo ed and exploi ed. LAB ha e shown
bene icial e ec s in ood-p oducing animals h ough a ious mechanisms. Acco ding o
he backg ound desc ibed p e iously, and, in pa icula , in a m animals, he EPS p oduced
by LAB may be pa o he mechanisms by which bac e ia gene a e some o he bene icial
e ec s in animals. EPS can enhance he iabili y o LAB in he gas oin es inal ac , bo h
due o i s esis ance o gas oin es inal condi ions, and by inc easing i s pe manence in
he in es ine (by bio ilm o ma ion, hey can inc ease in e ac ion wi h he cells o he
in es ine—bu in some cases hey dec ease i because hey in e ac wi h adhesion ac o s).
The adminis a ion o bo h EPS-p oducing s ains and pu i ied EPS can p e en a i ely
o he apeu ically p o ec hos animals om pa hogen in ec ions, inc ease he immune
esponse and imp o e eed con e sion, which esul s in lowe mo bidi y and mo ali y,
and g ea e eed e iciency, he eby gene a ing economic bene i s on a ms. In u n, hese
bene i s a e added o he ac ha he use o EPS, o s ains ha p oduce, hem can eplace
an imic obial d ugs.
In ensi e animal husband y has inc eased he use o an ibio ics bo h p e en a i ely
and he apeu ically. The use o al e na i e agen s o an ibio ics should be encou aged and
a wide ange o ools should be a ailable o p e en an ibio ic esis ance. Cu en ly he
addi ion o an ibio ics in o eeds u s o ood-p oducing animals is a global end, and hei
use p oduces an inc ease in an imic obial esis ance, which can be ansmi ed h oughou
he ood chain and gene a e heal h p oblems in consume s. The sea ch o EPS-p oducing
LAB, as well as he elucida ion o he mechanisms o ac ion o EPS in ood-p oducing
animals, will inc ease he quan i y o EPS, o p oducing s ains a ailable o adminis a ion
on a ms, as well as he knowledge o he expec ed bene icial e ec s, and will enhance he
a ailabili y o al e na i e s a egies o he use o an imic obials.
Au ho Con ibu ions:
W i ing—o iginal d a p epa a ion, M.L.W., A.M.H.-A., L.P.S. and L.S.F.;
w i ing— e iew and edi ing, M.L.W., M.T.D., M.J.R. and P.L.; in es iga ion and igu e design, M.L.W.,
A.M.H.-A. and L.P.S.; concep ualiza ion, M.L.W., P.L. and M.T.D.; supe ision, L.S.F.; p ojec adminis-
a ion and unding acquisi ion, P.L. and L.S.F. All au ho s ha e ead and ag eed o he published
e sion o he manusc ip .
Funding:
The p esen wo k was suppo ed by he Spanish Minis y o Science, Inno a ion and
Uni e si ies (g an RTI2018-097114-B-I00), by he Uni e si y o he Basque Coun y (GIU19/014)
and by CONICET (P oyec o PUE 058), Na ional Agency o Scien i ic and Technological P omo ion
(P oyec o PICT 2016-3495) and Uni e sidad Nacional del Li o al (P oyec o CAI+D 50120150100152LI,
CAI+D 50120150100151LI, CAI+D 50620190100152LI and CAI+D O ien ado 2016 2-14).
Ins i u ional Re iew Boa d S a emen : No applicable.
In o med Consen S a emen : No applicable.
Da a A ailabili y S a emen : No applicable.
Acknowledgmen s: We hank S ephen Elson o he c i ical eading o he manusc ip .
Con lic s o In e es : The au ho s decla e no con lic o in e es .
Foods 2022,11, 1284 25 o 34
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