9/13/2017 Disease prevention in Shrimp | Nutriad | Animal Feed Additives

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Disease prevention in Shrimp

Penaeid shrimp production is under continuous threat by bacterial and particularly viral infections
which have caused disastrous collapses of the industry in all major shrimp producing countries.

Disease problems in shrimp production are complex and often still poorly understood. Regulations,
consumer demands and sustainable management strategies restrict the number of drugs available
to treat pathogens. Vaccines are likely to be ine꺏㜭ective in crustaceans, which lack a speci㏺c immune
system similar to that of vertebrates.

Therefore, shrimp producers must consider the seed stock quality, husbandry procedures and
healthy nutrition as the major tools to control disease. The current article reports on recent progress
in the development of feed additives capable of reducing the impact of diseases on productivity and
pro㏺tability in shrimp farming.

Diseases are number one threat

The production of crustaceans has shown an average annual growth rate of 18% over the period 1970-2008,
which by far exceeds growth for all other aquaculture species (FAO, 2010). World shrimp aquaculture is
producing now well over 4 million MT of shrimp (Valderrama, 2011). This rapid increase in crustacean
production largely re醡罯ects the dramatic increase in white leg shrimp culture in China, Thailand, Vietnam and
Indonesia since 2000. Despite this apparent success story in terms of production expansion, shrimp
production in many regions continues to su꺏㜭er important economic losses due to the impact of a wide
variety of diseases.

Recent events illustrate the impact of disease outbreaks on shrimp production in major producing
countries. The white spot syndrome virus (WSSV), one of the main causes of the stagnating shrimp industry
in the nineties, is signi㏺cantly a꺏㜭ecting shrimp production in recent years in Mexico and Brazil. Early
Mortality Syndrome or Acute Hepatopancreatic Necrosis Disease (EMS/AHPND), is presently disrupting
production in the three major shrimp producing countries China, Thailand and Vietnam. EMS was ㏺rst
reported in China in 2009, it has spread to Vietnam, Malaysia and Thailand, and now causes annual losses of
billions of USD.

EMS outbreaks typically occur within the ㏺rst 30 days after stocking a newly prepared shrimp pond, and
mortality can exceed 70%. EMS is caused by a bacterial agent, which is transmitted orally, colonizes the
shrimp gastrointestinal tract and produces a toxin that causes tissue destruction and dysfunction of the
shrimp digestive organ known as the hepatopancreas.  The EMS/AHPND pathogen consist of a number of
unique strains of a relatively common bacterium, Vibrio parahaemolyticus, which have acquired the
capabilities to release the potent toxin.

Traditional approaches to boost shrimp health through the feed

A traditional approach to reduce the impact of shrimp diseases consists of increasing the level of key
nutrients a꺏㜭ecting the health and immunology of shrimp, including vitamin C and E, phospholipids, essential
fatty acids, trace minerals and carotenoids.

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These “booster feeds” are often supplemented with immunostimulants, mostly derived from the cell
envelope of micro-organisms, such as polysaccharides, lipoproteins, and lipopolysaccharides. The
continuous use of immunostimulants is generally discouraged due to the risks for over-stimulation of the
immune defense system.

Alternating on/o꺏㜭 regimes for feed additives is often impractical in farm operations. Encouraging results to
improve disease resistance have been obtained by the continuous use of health enhancing booster feeds
based on the selection of the appropriate immunostimulants in combination with a balanced nutritional
supply of key nutrients to support the enhancement of the immune system (Table 1).

However, the e꾌➪cacy of various commercially available immunostimulants to improve stress and/or disease
resistance of ㏺sh and shrimp strongly depends on the type of the product and on the supply of adjuvant
nutrients that are essential to support the buildup of the immune system.

Table 1: E꺏㜭ect of booster feed on production parameters in a farm in NE Brazil during episode of increased
disease incidence due to a combination of intensive rains and increased incidence of infectious myonecrosis
virus (IMNV) and necrotising hepatopancreatitis (NHP). Booster feed based on enhanced nutritional
speci㏺cations and supplementation of an immunomodulator (AQUASTIM S, Nutriad) versus standard feed.

(https://nutriad.com/wp-
content/uploads/2015/01/Article-Disease-Prevention-in-Shrimp-Figure-1.jpg)

Novel approaches (1): boosting the nutritional status and lipid reserves of the hepatopancreas

Shrimps do not tolerate high levels of dietary fat very well. A number of studies show reduced growth at
levels above 10% of dietary lipid. Nevertheless, quality and quantity of dietary lipids play a primordial role in
growth and health of shrimp. Shrimp have no or very limited capacity to biosynthesize a number of lipid
molecules which are essential for normal growth, including cholesterol, highly unsaturated fatty acids and
phospholipids. Fishmeal and ㏺sh oil are often the most important sources of cholesterol and HUFA in the
diet. Increasing cost of these marine ingredients has forced formulators to reduce dietary speci㏺cations for
these essential lipids.

Although these nutrient levels may not show signi㏺cant di꺏㜭erences on growth performance in feeding trials
under controlled conditions, they may become critical for maintaining health and immune defenses under
disease challenges and 醡罯uctuating ambient conditions encountered in production. Furthermore, the energy
status of shrimp is largely determined by its lipid reserves deposited in the hepatopancreas which functions
both as a digestive gland as well as a storage depot for energy. Therefore, farmers routinely look at squash
preparates to evaluate the nutritional status of the hepatopancreas, with ample lipid reserves being an
indicator of better resistance to stress and disease challenges.

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(https://nutriad.com/wp-content/uploads/2015/01/Article-Disease-
Prevention-in-Shrimp-Fig2.jpg)

Fig. 2: The hepatopancreas is the main organ of the shrimp’s digestive system responsible for digestion,
absorption and storage of nutrients. Esophagus (E), gastric mill (GM), hepatopancreas (HP), mid gut (MG),
hind gut (HG), and anus (A).

Lipid digestion in shrimp occurs for a big proportion intracellular in the hepatopancreas epithelium from
where it is transported to the target organs via the haemolymphe under the form of lipoproteins (Fig. 2).
The formation and absorption of lipid micelles from the lumen of the hepatopancreas tubuli is therefore a
limiting step in the lipid digestive process.

Digestibility enhancers based on natural emulsifying agents, selected for their compatibility with the
shrimp’s digestive system, have shown to be capable of complementing the process of emulsi㏺cation and
absorption of dietary fats in the hepatopancreas (Coutteau et al., 2012). This in turn improves the e꾌➪ciency
of shrimp to use fats as essential components and as source of energy for growth and surviving episodes of
stress or disease pressure.

The enhancement of the lipid reserves in the hepatopancreas of white shrimp Penaeus indicus as a result of
the supplementation of a digestibility enhancing additive was demonstrated recently by van de Braak et al.
(2012). Histological analyses showed a three-fold increase of the percentage of shrimp with a high degree of
lipid vacuolization in the hepatopancreas after supplementing the feed additive during one month (Fig. 3, 4).

The results of a parallel pond study indicated 2% higher average body weight (ABW), 4% higher survival, and
6% higher biomass for the treatment ponds. However, removal of outliers for survival from control and
treatment set showed 8% higher ABW, 12% higher survival, and 23% higher biomass.

(https://nutriad.com/wp-content/uploads/2015/01/Article-Disease-
Prevention-in-Shrimp-Fig3.jpg)

Fig. 3. E꺏㜭ect of the supplementation of a digestibility enhancing additive (Aquagest®S, Nutriad) on the
degree of lipid vacuolization in the hepatopancreas of shrimp fed the di꺏㜭erent feeds during 30 days (van de
Braak et al., 2012).

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(https://nutriad.com/wp-content/uploads/2015/01/Article-Disease-Prevention-in-Shrimp-Fig4.jpg)

Fig. 4: Histological determination of the degree of lipid vacuolization of the hepatopancreas in shrimp
Penaeus indicus, showing  a high (left picture) and low (right picture) level of lipid vacuolization (100x
magni㏺cation; van de Braak et al., 2012)

Novel approaches (2): Quorum Sensing technology

Quorum Sensing (QS) is a form of bacterial communication. Over the last decade, many bacterial species
have been documented to be able to produce and secrete small signaling molecules, such as acyl
homoserine lactones or certain oligopeptides, which can be detected by adjacent bacteria of the same or of
distinct species.  When population density raises, these molecules will accumulate in the extracellular
environment, thereby providing a means for bacteria to quantitatively monitor the presence of other
bacteria. These signaling molecules will, when reaching a certain threshold concentration, initiate
intrabacterial signaling that culminates in the activation of speci㏺c genes. QS communication is therefore
used by bacteria to synchronize gene expression alterations and coordinate biochemical responses within
the entire population.

In most pathogenic bacteria from which the QS system has been studied, QS has been associated with
pathogenicity, such as bio㏺lm formation and the production of proteases, invasion factors or other
virulence factors (Defoirdt, et al., 2011). In recent years, research focusing on ways to disturb QS signaling
(also called quorum quenching) is therefore gaining particular interest (Fig. 5). This is especially true in the
㏺eld of human medicine, where QS-inhibitors are investigated as potential alternatives to antibiotics in
tackling pathogenic bacterial infections (Sintim et al., 2010). Interestingly, chances that bacteria build up
resistance against QS disruptors are predicted to be low, giving that the selective pressure against these in
se non-lethal molecules is limited. This stands in stark contrast with what is seen with conventional
antibiotics (Defoirdt et al., 2010).

(https://nutriad.com/wp-content/uploads/2015/01/Article-
Disease-Prevention-in-Shrimp-Fig5.jpg)

Fig. 5: Quorum Sensing (QS), an innovative mechanism to tackle pathogenicity

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Initial studies of quorum sensing in aquaculture organisms are very limited but point out exciting results.
Halogenated furanones isolated from red marine algae, for example, have been demonstrated to reduce
QS-regulated gene expression in Vibrio and to protect ㏺sh and shrimp from vibriosis (Rasch et al., 2004;
Defoirdt et al., 2006).

At the Nutriad Technology Center, QS technology is being applied in a novel generation of natural feed
additives capable of modulating gut micro 醡罯ora. Compounds are tested for their capacity to inhibit QS-
signaling using an array of genetically modi㏺ed bacterial biosensors and QS-dependent infection protocols
in simple model organisms. Using these sensitive assays, potent QS modulators, able to shut down QS
signaling at concentrations far below the minimum inhibitory concentration, are being identi㏺ed.

Di꺏㜭erent QS quenching activities are selected for agriculture and aquaculture species based on screening
work using speci㏺c bacterial biosensors and model organisms. Synergistic blends of di꺏㜭erent natural
compounds resulted to be extremely e꾌➪cient in QS quenching activity against Vibrio harveyi signaling (Fig.
6).

(https://nutriad.com/wp-
content/uploads/2015/01/Article-Disease-Prevention-in-Shrimp-Fig6.jpg.png)

Fig. 6: Dose – response of a synergistic blend of botanical compounds (SANACORE GM, Nutriad) on Quorum
Sensing signaling activity of Vibrio harveyi. Graphs show signaling activity in QS biosensor system Vibrio
harveyi BB170, relative to control, exposed to di꺏㜭erent dilutions of the product extract (Nutriad Technology
Center, inhouse results).

Putting QS inhibition into practice: e꺏㜭ect of optimizing gut health on productivity and economics of
semi-intensive shrimp farming

Shrimp are actively “grazing” on the substrate present in the pond bottom and water column, and therefore
highly exposed to exchanges of micro醡罯ora between the environment and the digestive system. This
increases the risk for the proliferation of an unfavorable gut micro醡罯ora or frequent destabilization of the
micro醡罯ora, which can a꺏㜭ect the optimal functioning of the digestive system.

Furthermore, the digestive system of shrimp is the main entry port for bacterial and viral infections, which
remain a major risk for the pro㏺tability of shrimp production.

Sustainable approaches to modulate the gut micro醡罯ora in farmed animals include the use of selected
bacteria to inoculate the gut (probiotics), speci㏺c nutrients promoting the development of selected bacterial
strains (prebiotics), and speci㏺c natural compounds (mostly derived from yeast and herbal extracts, so
called “phytobiotics”) capable of modulating the micro醡罯ora towards a favorable composition, favoring the
development of bene㏺cial bacteria and inhibiting potentially pathogenic micro-organisms. The latter
strategies have the advantage of being easily applicable at the feedmill on large volumes of feed and
avoiding major adaptations of the production protocols at the farm.

A synergistic blend of botanical extracts (Sanacore® GM, Nutriad) was originally selected for its
bacteriostatic and bactericidal properties against pathogenic and potentially pathogenic bacteria in vitro
using the disk di꺏㜭usion method. Furthermore, this synergistic blend has proven to be a powerful interrupter
of bacterial QS signaling at concentrations well below minimal inhibitory concentrations (see above),
allowing it to e꺏㜭ectively modulate the gut 醡罯ora towards a more favorable composition.

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The supplementation of Sanacore GM promoted growth signi㏺cantly in healthy shrimp growing under
controlled lab conditions; showing a remarkable 20% increase of weekly weight gain and 4% improvement
on food conversion (Coutteau et al., 2010). The e꺏㜭ect of this botanical product showing combined activities
in QS inhibition and bactericidal action against a wide range of pathogenic bacteria was veri㏺ed in a semi-
intensive shrimp farm in Panama.

The second production season in Panama, stocked in aug-sept, is characterized by unstable climatological
conditions, resulting in strong temperature 醡罯uctuations which in turn a꺏㜭ect shrimp growth and increase the
impact of outbreaks of white spot virus (WSSV).

The ㏺rst production cycle, seeded in jan-april, provides more suitable growth conditions and generally
results in better survival and productivity. Two treatments were compared which only di꺏㜭ered with regard
to the supplementation or not of the phytobiotic growth promoter (Sanacore® GM) to the standard feed
used at the farm.

The supplementation of the botanical feed additive drastically improved survival, amounting to a relative
increase with 24% and 18% compared to the control group for the dry and wet cycle, respectively (Fig. 7).
Natural White Spot Disease outbreaks were observed during shrimp farming in both treatments under
similar frequency and severity; WSSV virus was con㏺rmed by immuno-chromatography and nested-PCR
tests.

The presence of a synergistic blend of phytobiotics provided an array of antimicrobial activities in the
shrimp’s digestive system. This o꺏㜭ered additional protection against co-infections with opportunistic
bacteria such as vibriosis, often the major cause of mortality in WSSV-infected shrimp (Phuoc et al., 2009).

The evaluation in the second cycle on 8 replicates per treatment allowed a good evaluation of variability
among ponds for the di꺏㜭erent production parameters. The addition of the phytobiotic reduced drastically
the variability of production results among ponds fed the same feed (average coe꾌➪cient of variation
between ponds for the 6 production parameters : control 18% versus Sanacore group 10%; Table 2). This
further indicated the importance of increased control of gut micro醡罯ora on the reproducibility of production
in semi-intensive pond environments.

(https://nutriad.com/wp-content/uploads/2015/01/Article-
Disease-Prevention-in-Shrimp-Fig7.jpg.png.jpg)

Fig. 7: Survival percentage at harvest for control ponds and treatment ponds receiving the phytobiotic
supplement in two production cycles in semi-intensive production of white shrimp L. vannamei (average and
standard deviation of 8 and 5 replicate ponds of 3ha per treatment, respectively; data from Vaca et al., 2010,
2011).

Table 2: Production results for P. vannamei in Panama during the second production cycle for control ponds
and treatment ponds receiving a phytobiotic supplement after 141 days of culture (average and standard
deviation of 8 replicate ponds of 3 ha per treatment).

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(https://nutriad.com/wp-content/uploads/2015/01/Article-Disease-Prevention-in-Shrimp-Table2.jpg)

Authors:
Peter Coutteau, PhD & Tim Goossens, PhD
Nutriad

Literature Cited:
Available from the authors

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