Plant based health promoters for secure shrimp farming, Using seaweed to prevent Vibrio

1. PhotofromtheInternationalAquafeedphotocompetition-photographer:©MatiNitibhon
36 | INTERNATIONAL AQUAFEED | January-February 2015
EXPERT T●PIC
Welcome to Expert Topic. Each issue will take an in-depth look
at a particular species and how its feed is managed.
SHRIMP
EXPERT TOPIC

2. The evolution
of modern
shrimp farming
by Malachi Stone, International
Aquafeed
A
lthough marine shrimp farming
has been practised in many
Asian countries for at least a
hundred years, it is only in the
past decade or so that it has really become
an economically important industry. In the
past, shrimp were viewed only as a second-
ary crop. Generally, fry were accidentally
washed by the tide into coastal paddy fields
and brackish fishponds. Trapped there,
they were simply allowed to grow to a
decent size then captured and sold. The
supply of fry would depend entirely on
fluctuations in the wild population. No
efforts were made to control predators or
competitors for food and space, and there
was no artificial feeding system. The young
shrimp were thus left to fend entirely for
themselves. Even the water they were living
in was usually too shallow to protect them
adequately from freak changes in weather
conditions. For all these reasons the yields
were somewhat haphazard. Even in a good
year, a farmer could only expect to harvest
between 100 and 300kg per hectare.
Then the market changed. Farmers real-
ised that the shrimp in their paddy fields
were beginning to sell for more than the rice
itself. So they simply converted their fields
and fishponds into shrimp farms.
Where used, modern shrimp farming
techniques have addressed many of the fail-
ings of the traditional system. Wherever pos-
sible, the farmer has taken active control of
the situation rather than leaving it to chance.
There is a greater density of shrimp in the
pools, because more seawater – and thus,
more fry - has been pumped in. The pools
have been dug deeper, providing a more
constant microclimate: the more water over
the shrimps’ heads, the greater the protec-
tion from environmental fluctuations.
One problem remains, however.
Expansion of the industry is still constrained
by an inconsistent supply of fry, which still
depends on captures from wild stocks. So far
only one species of shrimp, Penaeus chinensis,
is able to complete its entire breeding cycle
in captivity.
Shrimp in culture start out feeding on the
algae and aquatic plants that occur naturally
in their pools. However, as they get larger a
supplement is usually needed. Like lobsters,
they are often fed on trash fish, often mixed
with rice bran. Other feeds vary from: crushed
snails, mussel and clam meat, household
leftovers and even large pieces of cowhide
suspended in the water for them to feed on.
January-February 2015 | INTERNATIONAL AQUAFEED | 37
EXPERT T●PIC
1
2

3. D
r Eckel, a leading supplier for research-based functional
feed additives is about to launch its Shrimp product line
in Thailand and South-East Asia in 2015. New research
results from Thailand corroborate the strength of these
future standards in plant-based health promoters.
Early Mortality Syndrome (EMS) is still a big problem in shrimp
production in South-East Asia as well as in Latin America. EMS experts
know that classical approaches like antibiotics and chemical disinfection
are no solution:
“Disinfectants are only good for a couple of minutes, so they are
not the answer. Antibiotics are not the answer, either. The vibrios are
there, and we have to keep them in check with balanced systems,”
says Scott Horton.
Centrepieces of such balanced systems are farms with strictly man-
aged biosecurity and well-considered genetic resources and breeding
practices. Another task is to strengthen the natural defence mechanism
of the animals in order to cope with any kind of disease pressure.
Functional feed additives are one way to fulfill this task.
Demand for natural solutions
For several years customers and retailers in Europe have become
more and more sensitive to the abundance of antibiotic drugs usage
in animal production systems. Consumers and policymakers oppose
unjustified use of antibiotic drugs either from an individual health
perspective or from a global sustainability perspective. Shrimp farming
enterprises who want to keep pace with that development have to
change their practices and have to look for alternative ways to cope
with disease pressure in intensive shrimp farming systems.
R&D efforts come to fruition
Dr Eckel was one of the first European feed additive producers who
expanded its business to aquaculture and aquafeed industries. The suc-
cess stories in livestock feed was encouragement to transfer the natural
nutraceutical concepts to aquafeed applications. Hence, the target was
to develop a functional feed additive that has positive effects on growth
and immune defence of Pacific White Shrimp.
After two years of research and development Dr Eckel is proud to
reap the fruits of its labours. In 2014, the Dr Eckel research partners
of the Aquaculture Business Research Centre at Kasetsart University
in Bangkok reported a breakthrough. The tested phytogenic feed addi-
tives proved to have positive effects on growth, survival and immune
response of shrimp.
In a series of tank trial experiments at Kasetsart University the
effects of dietary supplementation of phytogenic feed additives was
evaluated at different inclusion rates (400 ppm and 800 ppm) on
growth, survival, intestinal bacteria, immune responses and tolerance to
Vibrio parahaemolyticus infection in Pacific White Shrimp (Litopenaeus
vannamei). Postlarvae 12 (PL12) were kept in 500-litre tanks with sea-
water of 20-25 ppt salinity and controlled water temperature of 29+
1°C. PL12 were stocked at a density of 50 PL per tank (100 PL/m2).
In the growth experiment shrimp were fed for 60 days, the challenge
experiment lasted 30 days, with Vibrio being added before stocking
and at day 14 of the experiment. Each feeding group was carried out
with four replicates.
Increased body weight gain
The Dr Eckel feed additive enhanced the growth rate of shrimp in
laboratory condition in a dose-dependent manner. After 60 days of
dietary administration, shrimp with 800 ppm inclusion rate showed the
highest average body weight of 3.48 ± 0.18 g, followed by the 400 ppm
group with 3.42 ± 0.22 g. Shrimp from the control group showed the
lowest average body weight of 2.64 ± 0.43 g. These differences were
statistically different at a p-level of five percent. The researchers pre-
sume that the feed additive improved digestibility of nutrients leading
to higher feed efficiency and faster body weight gain. This hypothesis
is underpinned by the improved feed conversion ratio observed in the
treatment groups.
Better survival rates after Vibrio challenge
Survival rates did not differ between groups in the growth experi-
ments. However, in the challenge experiment shrimp from the group
fed with Dr Eckel feed additives at 800 ppm and at 400 ppm had
significantly higher survival rates (78 percent and 67 percent respec-
tively) compared to shrimp in the control group (64 percent) when
challenged with a virulent strain of Vibrio parahaemolyticus.
Researchers marvel at immunological effects
During the experiments, special emphasis was put on immunologi-
Figure 1: Protective feed additives are a building block of
healthy shrimp farming
1
by Tilman Wilke, Susanne Kirwan (Dr. Eckel GmbH, Niederzissen, Germany), and Niti Chuchird,
Hataitip Niyamosatha (Aquaculture Business Research Center, Kasetsart University, Bangkok, Thailand)
38 | INTERNATIONAL AQUAFEED | January-February 2015
EXPERT T●PIC

4. cal effects as one of the target mechanism of the novel feed additive.
The tested feed additive improved shrimp immune response, which
led to higher survival rates in the challenge experiment. In the course
of their immunological studies the researchers measured the ratio
of hemocytes cells that do phagocytosis to the total number of
hemocytes. Phagocytosis is the central and terminal mechanism of the
immune system to seek and destroy pathogenic bacteria or infected
cells. The share of hemocytes that perform phagocytosis was signifi-
cantly larger in the treatment group (23 percent) than in the control
group (17 percent). Additionally shrimp in the treatment group had
twice as much hemocytes compared to shrimp of the control group
(4.4 x 106 versus 1.9 x 106 cells / ml).
When the researchers examined the hepatopancreas of the chal-
lenged animals they discovered a spectacular histopathological pattern:
Shrimp from the treatment group had less cell necrosis in the hepato-
pancreas compared to other groups (Figure 2). That means that the
phytogenic feed additive from Dr. Eckel took a cell-protective effect
in the hepatopancreas of shrimp.
Modes of action under investigation
Although the biological effects are proven with scientific methods,
the molecular pathway of these effects is still under investigation. The
active ingredients in the feed additives are plant substances and plant
extracts that feature highest levels of flavonoids and polyphenols.
These substances are highly potent nutraceuticals that act as anti-
oxidant, free radical scavenger, anti-inflammatory agent and immune
system modulators.
Implications for shrimp farmers
and shrimp feed producers
With the new shrimp-adapted versions of his natural phytogenic
feed additives Dr Eckel delivers powerful tools that assure high pro-
duction and low mortality in intensive shrimp production. The proven
effects on growth, survival rate and immune response will make them
building blocks of a sustainable and balanced shrimp farming system.
Feed mills’ purchasers will be glad to hear that these feed additives
from Dr Eckel are heat-stable and can be ground or even pulverised,
what makes them suitable for extrusion and micro diet formulation.
And they work in low concentrations – leaving enough space for
nutritional ingredients.
LEIBER® BETA-S
EXCELLENT FOR FISH
REAL
BREWERS‘
YEAST
Made in German
y
•MadeinGermany•M
adeinGermany•Made
inGermany•MadeinGe
rm
any •
Leiber GmbH
Hafenstraße 24
49565 Bramsche
Germany
Tel.+49 (0)5461 9303-0
Fax +49 (0)5461 9303-29
www.leibergmbh.de
info@leibergmbh.de
VIV ASIA 11–13 Mar 2015, Bangkok, Thailand
Meet us: Welcome Hall, Stand A005
For strong, immunocompetent fish:
Improvement of the cellular &
humoral defence mechanisms
Support of immunological competence
in larval and juvenile stages
Improvement of feed conversion
Beta-S Viking ProBTRBierhefe®
Bierhefe
W60
MTMannan®
Figure 2: Histology of the hepatopancreas of control shrimps
showing 30 percent cell necrosis (left) compared to the
hepatopancreas of shrimp of group fed 400ppm of Dr. Eckel
phytogenic feed additive showing 5 percent cell necrosis (right).
Magnification 500x, Hematoxylin and eosin stain. (Copyright:
Niti Churchid, Kasetsart University)
January-February 2015 | INTERNATIONAL AQUAFEED | 39
EXPERT T●PIC

5. 2
W
ith unique expertise in
the use of clay and algae,
Olmix has developed
a new product aimed at
the improvement of shrimp performance
through increased digestive enzyme activity
and better digestive balance vital in prevent-
ing Vibrio. The solution lies in improving
gut health.
Shrimp farming started to develop in
the 1970s. In 2012, world shrimp produc-
tion was over 4 million tonnes and more
than 50 percent of the shrimp eaten in the
world today comes from aquaculture. South
East Asia and China represent the largest
and the most productive shrimp production
areas, accounting for 80 percent of world
production. Although shrimp production has
boomed over the last few
decades, farmers have to
face a variety of issues to
ensure their production.
Shrimp are very sensitive
animals and many disease
outbreaks have occurred
in the past, such as white
spot viral disease in 1994-
1995 in south East Asia,
and some are still ongoing,
such as Early Mortality in
Shrimp syndrome, hitting
stocks across South East
Asia since 2010.
One of the most
important of these dis-
eases is Vibriosis, which
kills shrimp at all stages
of the production cycle.
More than ten Vibrio spe-
cies have been reported
pathogenic for shrimp.
Though Vibrio bacteria are part of the natural
flora and culture environment of shrimp,
Vibriosis can occur in a context of imbalanced
environment and may cause total mortality
of the reared shrimp. During the last two
decades, mass mortality incidents in growout
ponds and hatcheries due to Vibrio were
widely reported. Among the different Vibrio
species, Vibrio harveyi is considered one of
the most important shrimp pathogens.
With very good management practices,
it is possible to limit the Vibrio problem. In
order to have complete protection against
the pathogen, it is important to find ways to
prevent shrimp being contaminated by Vibrio.
Antibiotics and chemotherapy are often used
to manage disease outbreaks. However, these
methods have limits, such as environmental
hazards or the spread of antibiotic-resistant
bacteria. Another way to prevent Vibrio
disease is improved gut health. The gut is one
of the most important conduits used by Vibrio
to infect shrimp. It is possible to favour the
natural defences of the gut by preserving its
natural balance to avoid Vibrio development
and toxicity.
While there is scientific evidence show-
ing the benefits of clays in the prevention
or treatment of digestive problems and in
the protection of the gut mucosa, much less
is known about their capacity to improve
feed efficiency. Yet the improvement of the
digestibility of feed is an integral property of
clays. The mechanisms involved are thought
to be multiple (Reichardt, 2008). The domi-
nant hypothesis described in the literature
is that clays slow down the transit of feed
in the intestine, so the time for digestion is
by Jean Peignon, aqua technical service, Olmix, Vietnam
Growth performance Control MFeed+ 0.1
percent
Variation over
control
MFeed+ 0.2
percent
Variation over
control
Average initial weight, g/ind 6.32 ± 0.13 6.32 ± 0.13 / 6.32 ± 0.13 /
Average final weight, g/ind 12.4 ± 0.8 13.09 ± 1.1 +5.6% 13.44 ± 0.1 +8.4%
Specific growth rate, percent/d 1.22 ± 0.1 1.31 ± 0.1 +7.4% 1.34 ± 0.1 +9.8%
Total feed consumption, g/ind 8.66 ± 0.64 8.21 ± 0.92 / 8.52 ± 0.77 /
Feed Conversion Ratio 1.42 ± 0.1 1.22* ± 0.09 -14.1% 1.23* ± 0.1 -13.4 percent
* p-value < 0.05
Health performance Control MFeed+ 0.1
percent
Variation over
control
MFeed+ 0.2
percent
Variation over
control
Vibrio at 60 days, x104 CFU/g - - - - -
In hepatopancreas 3.07 ± 0.39 2.83 ± 0.31 -7.8% 1.17** ± 0.13 -61.9%
In the intestine 1.67 ± 0.30 1.58 ± 0.25 -5.4% 1.13 ± 0.12 -32.3%
Survival rate, percent 67.2
± 4.38
78.4**
± 4.56
+16.7% 82.4** ± 3.58 +22.6%
** p-value < 0.01
40 | INTERNATIONAL AQUAFEED | January-February 2015
EXPERT T●PIC

6. increased, hence a better digestibility of feed
and increased nutrient uptake.
Nevertheless, it seems that the action
of clays to enhance feed digestion in the
intestine also involves other mechanisms.
Reichardt (2008) and Habold et al (2009)
both report the ability of clays to favor the
contact between enzymes and nutrients and
therefore to improve the rate of digestion
of the feed. Indeed, digestive enzymes need
to be in contact with their substrate in order
for hydrolysis to occur. The physico-chemical
interactions of the enzymes with clay particles
seem to enhance the contact between the
digestive enzymes and the feed, making clays
a good supporting matrix for enzymes and
acting as a meeting point for them to be in
contact with their substrate. Indeed, Cabezas
et al (1991) demonstrated that clay-enzyme
complexes are formed at enteric pH values.
These active stable complexes are resistant
to proteolysis and increase the amount of
active digestive enzymes in the intestine, thus
improving nutrient digestibility.
In the same way, Habold et al (2009)
observed higher pancreatic lipase activity in
rats supplemented with Kaolinite; Xia et al
(2004) showed an increase in small intestinal
digestive enzyme activity in broilers supple-
mented with Montmorillonite; and Paolo et
al (1999) observed an increase in protein and
energy retention coefficients for growing pigs
supplemented with clay. Some studies also
suggest that the increased activity of enzymes
in contact with clay comes not only from their
stabilisation, but also from the presence of
cofactors in the clay (Reichardt, 2008; Habold
et al, 2009). Cofactors are defined as thermo-
stable non-protein compounds that form the
active portion of an enzyme system. In other
words, cofactors are helper molecules required
for enzymes to be active. They can be organic
January-February 2015 | INTERNATIONAL AQUAFEED | 41
EXPERT T●PIC
After adding VIUSID® aqua to each feed intake throughout the
production process, the following results were:
A specific product to improve
productivity of aquaculture
www.catalysisagrovet.com

7. or inorganic, most commonly vitamins in the
first case and metallic ions in the latter.
Clays are layered mineral materials, com-
posed of a succession of aluminum and
silicium based sheets, the order of which
varies depending on the type of clay. In
Montmorillonite, several metallic ions replace
some aluminum and silicium ions in the struc-
ture. Known as the substitution phenom-
enon, this event provides montmorillonite
with part of its physico-chemical reactiv-
ity. Moreover, the presence of metallic ions
may contribute to the activation of some
enzymes, through their action of cofactors
(Niederhoffer, 2000). In this way, copper
is known to activate lipase and phospho-
lipase A (Jondreville et al, 2002) and zinc
is a required cofactor of carboxypetidase
(Williams, 1960), to mention only a couple
of examples.
The combination of the matrix support
provided by the clay and the cofactor effect
coming from the metallic ions present in its
structure can be referred to as biocatalysis:
the improvement of performance of a bio-
chemical reaction through the action of an
external compound, a biocatalyst. Due to a
large variety of clay minerals, one can imagine
that all clays do not have the same potential
for biocatalysis depending on their type, their
purity, their source or their treatment. As
such, clay structure can be modified and asso-
ciated with other materials in order to unlock
its biocatalytic properties. Such technology
has been developed by Olmix group (France)
in the course of its research conducted on
seaweeds and clays. The micronized form
allows a fine dispersion of the product in the
intestine, providing many sites of enzymatic
digestive reaction with more easily accessible
metal ions.
Moreover, it benefits from a synergy
between clay and seaweeds in the pro-
cess of biocatalysis, as seaweeds bring in
many diverse metallic ions, sometimes absent
in the feed, which are required cofactors
for the activation of several enzymes. This
unique combination of seaweeds and clay
makes it a unique tool to boost enzyme
activity through the action of biocatalysis.
MFeed+, the only product benefiting from
this new technology, has proven its effi-
cacy in several studies. In one, MFeed+ was
tested on shrimp by researchers at Kasetsart
University (Thailand). The aim of the study
was to evaluate the effect of MFeed+ feed
supplementation on digestive and growth
performance of Penaeus vannamei.
375 shrimp (6.3±0.2g weight) were dis-
tributed in fifteen 500 L glass tanks contain-
ing 25 shrimp each. After a 7-day period of
acclimatization during which all shrimp were
fed the basal diet, tanks were randomly allot-
ted to one of three treatments (5 replicates
per treatment): one control, fed the basal
diet and two MFeed+ groups, for which
the basal diet was supplemented with 0.1
percent or 0.2 percent MFeed+. Growth
performance parameters and mortality were
recorded during the 60 days of supplemen-
tation. Vibrio bacteria were counted in the
hepatopancreas and the intestine at 60 days
as a marker of digestive health. The shrimp
were fed three times a day to satisfaction.
The feed amount was adjusted daily based
on the feeding ability of the shrimp. Uneaten
feed was siphoned out of the tank 2 hours
after feeding. Water used in the experiment
was seawater, with salinity adjusted to 12-15
ppt. The water in the tanks was aerated with
air stone and exchanged every 2-3 days at
the rate of 10-30 percent volume depending
on its visible quality.
Feed Conversion Ratio was greatly
improved in groups receiving MFeed+. As a
consequence, average final weight and specific
growth rate of shrimp supplemented with
MFeed+ tended to be higher. Moreover, bet-
ter digestive performance helped to improve
the digestive status of the shrimp, as dem-
onstrated by a lower Vibrio count in the
hepatopancreas and the intestine and the
improved survival rate of the juveniles. This
study highlighted the potential of MFeed+ to
improve digestive and zootechnical perfor-
mance of shrimp.
42 | INTERNATIONAL AQUAFEED | January-February 2015
EXPERT T●PIC