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10
4
CFU g
-1
, but after
10 weeks feeding on
E. faecium
supplemented diets the total culturable counts increased to
10
7
CFU g
-1
compared to 10
6
CFU g
-1
in the control fed fish.
E.faecium
levels in the probiotic
fed fish accounted for over 70% (6.88 × 10
2
CFU g
-1
) of the epithelial mucosal populations
and nearly 90% (6.07 × 10
7
CFU g
-1
) of the digesta populations (Figure 8.1B). Re-population
of the GI tract with probiotics after antibiotic administration is likely to create competition
for potential pathogens re-entering the digestive tract and stabilize/reinforce the gut microbial
defensive barrier, and therefore it is suggested that this topic be further investigated.
Recently Avella
et al
. (2011) administered
E. faecium
IMC 511 to common sole (
Solea
solea
Linnaeus, 1758) larvae from the onset of the opening of the mouth (2 dph) to 50 dph.
At day 50,
Vibrio
counts were significantly lower than in the control group and LAB levels
were significantly higher in the probiotic groups at all time points (10, 30 and 50 dph). RAPD
analyses of isolates demonstrated that the
E. faecium
component of cultivable LAB popula-
tion increased in the probiotic groups with trial duration, peaking at 80-100% at 50 dph. The
probiotic was not detected in the control group at any time point. The modulation of GI micro-
biota was associated with a better growth and a beneficial effect on animal welfare supported
by lower levels of 70 kDa Heat Shock Protein.
All of the aforementioned studies have used culture-dependent approaches; to the authors'
knowledge only two studies have used a culture-independent approach to assess the effect of
enterococci probionts on the gut microbiota of fish (Sun
et al
. 2012a; Del'Duca
et al
. 2013).
Sun
et al
. (2012a) assessed the autochthonous microbiota in the fore-, mid- and hindgut of
juvenile grouper after the dietary administration of
E. faecium
MM4, previously isolated from
the whole intestine of juvenile grouper (Sun
et al
. 2009), at 10
8
cellsg
-1
for 60 days. DGGE
analysis revealed that the
E. faecium
MM4 successfully populated all three gut regions, of all
replicates, but was absent in the control fish. Subsequently, bacterial richness and diversity in
the midgut and hindgut were significantly elevated in the probiotic fed fish. Four OTUs (clos-
est known relatives identified as uncultured Betaproteobacterium (82% similarity), uncultured
Alphaproteobacterium (85%),
Comamonas
sp. MPI12 (90%) and
Dietzia psychralcaliphila
(94%)) were present only in the probiotic group. Del'Duca
et al
. (2013) observed that dietary
provision of
Enterococcus
sp. (
for 10 days. Viable counts after oxolinic acid administration were
<
10
6
bacteria g
-1
) elevated the total bacterial levels and entero-
cocci levels in the gut of tilapia. In addition, the application led to a reduction in the abundance
of putative
Pseudomonas fluorescens.
.
Taken together, these studies reveal that
E. faecium
strains possess the capabilities to pop-
ulate the gut, and modulate the gut microbiota, of rainbow trout, carp, grouper, European eel,
common sole and sheet fish.
>
8.3.4
Lactococcus
spp.
To the authors' knowledge the only lactococci strains which have been investigated with
regards to intestinal colonization and microbial modulation of fish
invivo
belong to the species
Lactococcus lactis
(Villamil
et al
. 2002; Balcázar
et al
. 2007a; 2007b; 2009; Perez-Sanchez
et al
. 2011; Mohapatra
et al
. 2012; Sun
et al
. 2012b; Table 8.2).
Villamil
et al
. (2002) reported that the
in vivo
adhesion to, and colonization of, the tur-
bot intestine by
Lc
.
lactis
was low. Despite demonstrating that
Lc. lactis
was able to adhere
to turbot intestinal mucus
in vitro
,afterthe
in vivo
experiment the bacteria were recovered
from only one fish (of five sampled, at 1.5 × 10
3
cells ml
-1
). It was suggested that the adher-
ence capabilities of
Lc. lactis
could have been reduced
in vivo
by environmental factors such
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