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2B (Chr. Hansen, Denmark) which contains
B. subtilis
(DSM 5750) and
B. licheniformis
(DSM 5749).
Bagheri
et al
. (2008) fed rainbow trout fry diets containing
Bacillus
levels ranging from ca.
5 × 10
8
to 6 × 10
9
CFU g
-1
for 2 months. This led to a significant increase of viable bacterial
counts in the intestine compared to the control group at the end of the trial. Total aerobic
bacterial counts were significantly different among treatments, suggesting that the indigenous
microbiota was affected by dietary probiont level. The percentage of probiotic
Bacillus
spp.
recovered from the intestine ranged from 65% to over 99% of the total viable culturable
counts. Changes in the relative proportions of the indigenous microbial groups were also
observed:
Pseudomonas
spp. and members of the Vibrionaceae seemed the least sensitive
as they remained the dominant members of the non-probiotic population. However, as the
probiotic supplementation level increased, Enterobacteriaceae and a group of unidentified
Gram-negative spp. were gradually phased out. Merrifield
et al
. (2010c) fed rainbow trout
B. subtilis
and
B. licheniformis
(5 × 10
7
CFU g
-1
) for 10 weeks and reported that
Bacillus
levels accounted for over 35% (1.42 × 10
3
CFU g
-1
) and 60% (1.17 × 10
7
CFU g
-1
)ofthe
culturable microbial population on the mucosa and in the digesta, respectively (Figure 8.1A).
In a second study Merrifield
et al
. (2010d) investigated the effects of
B. subtilis
and
B.
licheniformis
on the gut microbiota of rainbow trout pre-treated with an antibiotic. Viable
allochthonous counts after oxolinic acid administration were
10
4
CFU g
-1
but after 10
weeks feeding on probiotic supplemented diets (ca. 6 × 10
7
CFU g
-1
) total culturable levels
increased to 10
7
CFU g
-1
, compared to levels of 10
6
CFU g
-1
in the control fed group.
Bacillus
levels accounted for 75% (5.5 × 10
3
CFU g
-1
) of the culturable mucosal associated
populations and over 80% (2.6 × 10
7
CFU g
-1
) of the digesta populations (Figure 8.1B).
Although a common strategy in mammals (Madden
et al
. 2005), to the authors' knowledge
this is the first study in fish to incorporate the use of a probiotic after the use of antibiotics.
Antibiotics remain an important part of the arsenal of treatments for many fish bacterial
diseases and are used globally, and such applications can alter the gut microbiota in terms
of total viable numbers and/or diversity of populations (Austin and Al-Zahrani 1988; Lesel
et al
. 1989; Moffitt and Mobin 2006; Bakke-McKellep
et al.
2007; Liu
et al
. 2012; Romero
et al
. 2012). If reductions in the microbial levels, diversity or composition occur it could
open a niche leaving the GI tract susceptible to secondary potential pathogens. The study
of Merrifield
et al
. (2010d) demonstrates that probiotic supplementation after antimicrobial
treatment increases the number of viable bacteria within the digestive tract and associated
with the epithelial mucosa via repopulation with viable probionts. This is likely to create
competition for potential pathogens re-entering the digestive tract and stabilize/reinforce the
gut microbial defensive barrier after antibiotic treatment.
These studies demonstrate that dietary
B. licheniformis
and
B. subtilis
can survive transit
through the rainbow trout digestive tract and successfully populate the intestine as both tran-
sient and potentially resident populations providing that continual dietary supplementation is
applied. The evidence also suggests there is little strain specificity, as both commercial
B.
subtilis
and autochthonous (strains isolated from fish)
B. subtilis
strains have displayed high
intestinal recovery after dietary supplementation.
In addition to rainbow trout, it has been demonstrated that dietary
B. subtilis
can modulate
the gut microbiota of a number of live bearing ornamental fish species. Ghosh
et al
. (2008)
reported that intestinal
B. subtilis
levels significantly increased from being non-detectable to
>
<
log 6-7 CFU g
-1
in the gut of
B. subtilis
fed guppy (
Poecilia reticulate
), molly (
Poecilia
sphenops
), green swordtail (
Xiphophorus helleri
) and southern platyfish (
Xiphophorus
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