Sustainable Shrimp Farming with Beneficial Microbes
Introduction
Shrimp farming
is considered a major factor in bringing the blue revolution around the world. It
has grown several folds during the last two decades and transformed the
aquaculture sector into a multimillion-dollar industry. Disease outbreak is a nightmare for shrimp
farming and its management is a big challenge. Shrimp is highly prone to viruses
and bacterial pathogen hence intensive management is required to avoid the
outbreak of disease. Huge economic
losses have been observed in shrimp farming in India primarily due to virus pathogens
in addition to the bacterial pathogens as well. An estimate says that around
1000 crore INR was lost during 2006-08 due to disease in shrimp also affecting the
employment of 2.15 million man-days (http://epubs.icar.org.in/ejournal/index.php/FT/article/view/26607).
Epidemic significance between the diseases was formerly for White Spot Syndrome
Virus (WSSV), TSV and recently for bacterial disease such as Loose Shell
Syndrome (LSS) and white gut and slow growth syndrome and recently EHP too. Use
of chemicals and antibiotics are always discouraged in aquaculture due to
associated side effect such as antibiotic resistance, chemical residues, cost,
etc. Alternative of chemicals is always a welcome step for environment-friendly
sustainable shrimp farming. Application of microbes for successful shrimp
farming is now a day considered as the most effective approach and gaining
popularity among the aquaculturist.
The aquatic
system is considered as a home of the innumerable numbers of microbial diversity.
These microbes play a major role in the dynamics of the water ecosystem through
the recycling of nutrients. Further several microbes are identified as a
potential pathogen inhibitor and considered beneficial microbes for health
management in aquaculture. These microbes are found in the gills, intestine,
muscle, on the body surface of fish, shrimp, water, soil, etc. and include
bacteria, fungi, algae and protozoa. These microbes may be beneficial or
harmful but plays a significant role in aquaculture. The beneficial role played
by microbes in aquaculture includes a role in bioremediation, as gut
probiotics, as live food, food supplements, probiotics etc. The following
section will elaborate on some of the important aspects of the microbial
application for sustainable shrimp farming.
Probiotics
are regarded as a new generation disease management approach in aquaculture.
Now a day’s probiotics are used enormously in shrimp farming for disease
management. The term probiotic was derived from the Greek words “pro” and
“bios” means “for life,” (Gismondo et al., 1999) and introduced by Lilly
and Stillwell in 1965 as a modification of the original word “probiotika.”.
Fuller et al. (1989) distinct it as “alive microbial feed addition which
constructively marks the host animal by improving its intestinal microbial
balance”. Verschuere et al. (2000) further revealed it as “a live
microbial adjunct which has a valuable outcome on the host by modifying the
host-related or ambient microbial community by confirming improved use of the
feed or growing its nutritional value, by enhancing the host response towards
disease, or by improving the quality of its ambient environment”. Probiotic displays
its effectiveness through the different mechanisms as listed subsequent- (1)
Competitive exclusion – probiotic organism colonizes the gut thereby inhibiting
colonization of pathogenic bacteria. (2) Probiotic organisms produce certain
inhibitory substances which obstruct pathogenic organism. (3) Competition for
nutrients – probiotic organism utilizes the available nutrients and hence the same
is unavailable for the pathogens. (4) Substances produced by probiotics act as an
antagonist for the quorum sensing network of the pathogen. (5) Improved
immunity – augmented macrophage activity and antibody level. (6) Enhancement in
water quality. (7) Interface with phytoplankton. (8) Antiviral activity
(Zorriehzahra et al., 2016).
used in shrimp farming
Probiotics are mainly of two types
a) Gut
probiotics are included with feed to improve the useful microbial flora of the
gut and maintain gut health and also suppress the proliferation of pathogenic
organisms in the gut.
b) Water probiotics are basically added into
the water to maintain handsome water quality by reducing the level of the toxic compound such as ammonia, nitrate, sulphate, etc.
Feed
probiotic
Feed probiotics
are mixed with feed along with binders for better stabilization (Kolndadacha et
al., 2011). It is aimed to establish a balanced gastrointestinal microbial
community to improve digestive as well gut-associated immune system
response. Application of probiotics a supplemented feed is now a day common practice in shrimp farming. Probiotics,
including bacterial strains, yeast and extracted substances are generally
supplied by this method. Some probiotics that have been added in animal feed
include bacterial species, such as Lactobacillus spp., Enterococcus
faecium, Bifidobacterium, Thermophilum, Streptomyces spp.,
Micrococcus spp., Pseudomonas fluorescens, as well as yeast, such as Saccharomyces
cerevisiae.
Water
probiotic
Water probiotics
are applied directly to the culture pond to improve water quality (Boyd, C.E.,
1989) to provide stress free environment and in response improves the survival percentage
of animals (Moriarty et al.,
1998). The mode of action of water probiotics are actually based on bioagumentation
or biocontrol of toxic compounds present in water and also improvement of the
microbial ecology of the water and sediment (Rengpipat et al., 1998). Numerous biological yields,
such as live bacterial inoculum, enzyme preparations, and plant substrates
extracts have been used for water and soil quality improvement factors in
aquaculture ponds (Boyd C.E., 1989). Water probiotic tends to minimize the
level of the different toxic compounds such as TAN, organic load, nitrite etc.
and provide a healthy and environment to the culture organism.
Probiotics
application in Shrimp farming
Probiotics
are applied enormously in shrimp farming all around the world as a measure of
disease management and growth stimulator. Rengpipat et al. (1998)
isolated Bacillus S11 bacterium from
black tiger shrimp territories and supplementary with feed to Penaeus
monodon PL30 for 100 days in three formulas: fresh cells, fresh cells in
normal saline solution, and lyophilized form. PL30 exhibited no noteworthy the difference in growth, survival and external form of all three probiotic treatments
whereas major differences were detected between probiotic and control groups.
After stimulating shrimps with a pathogen, Vibrio
harveyi, by immersion for 10 days, all probiotic treatment groups had 100%
survival; whereas the control group had only 26% survival.
Soundarapandian et
al. (2008) reported higher average body weight, survival, production, and
low FCR as compared to control after the application of Super -PS probiotic in Penaeus monodon cultured pond. Further,
they also observed the prevalence of bacterial infection such as gill socking
and tail rot disease in control which was absent in treatment.
Wang et al.
(2010) detected that the usage of water probiotic Lactobacillus acidophilus, Rhodopseudomonas
palustris and Bacillus coagulans meaningfully
improve the final weight, daily weight gain and relative weight gain of the
shrimp compared to control. After 35 days, no significant difference in final
weight and relative weight gain were found. However, significantly higher daily
weight gain was observed with Bacillus
coagulans compared to Rhodopseudomonas
palustris. The immune response showed
a remarkable increase in PO activity in shrimp treated with Bacillus coagulans compared to Lactobacillus acidophilus and Rhodopseudomonas palustris. Javadi et
al. (2011) fed commercial probiotic (Protexin) incorporated diet to Penaeus indicus and found a significant
increase in weight and growth rate of shrimps at the end of the breeding
period. Sivakumar et al.
(2012) used Lactobacillus acidophilus
as a probiotic against Vibrio in Penaeus monodon juveniles. L. acidophilus inhibited the
proliferation of Vibrio parahaemolyticus,
Vibrio cholerae, Vibrio Harvey and Vibrio
alginolyticus. The effects of L.
acidophilus was verified by feeding through the feed for 30 days before and
after an immersion trial with V. alginolyticus. Survival was determined
after 10 days of the challenge. The result showed 20% final mortality in the
treated group and 86.7% in the control group.
Kumar et al.
(2013) compared the effects of feed probiotic Bacillus subtilis and L. rhamnosus on the growth performance of
shrimps. B. subtilis showed a better
effect than L. rhamnosus in terms of
growth performance. Further superoxide dismutase (SOD) and catalase activity of
the haemolymph in shrimps fed with B.
subtilis showed enhanced activity as compared to L. rhamnosus. Elumalai et al. (2013) reported improved
growth performance, survival and beneficial bacterial community of P. monodon upon application of mixed
Bacillus probiotics. Sandeepa et al. (2015) used Lactobacillus sp. as probiotics at three different concentrations
in shrimp basal diets. The results revealed that lower concentration got better
growth performance than those fed with basal diet. A significant increase in
glucose, protein and triglyceride was reported in shrimp fed with lower, medium
and higher level probiotics respectively. Yuvaraj et al. (2015) reported
better growth and disease control in a zero water exchange culture system of
shrimp Litopenaeus vannamei after the
application of probiotics and/or enzymes as compared to control.
of Probiotics on Vibrio in Shrimp Aquaculture
Vibrio spp. occurs obviously
in aquatic environments and is one of the most major bacteria in shrimp farming
(Vandenberghe et al., 2003). Some of the pathogenic Vibrio species have also been detailed as the causative agents of
shrimp infections (Goarant et al., 1999). When high temperatures and salinity
condition,various species such as Vibrio
parahaemolyticus predominate in shrimp pond (Williams and LaRock, 1985).
Noriega-Orozco et al. (2007) reported that the pond conditions in shrimp
farms may support the survival and growth of high salt-tolerant pathogenic Vibrio species, such as V. parahaemolyticus, V. harveyi, V. alginolyticus. Wang et al. (2005) reported a
decrease in vibrio count after the application of probiotics. Far et al.
(2009) informed abridged Vibrio spp
count in the digestive tract in L.
vannamei after application of the higher level of B. subtilis. Vieira et al. (2010) conveyed reduced total
bacterial count and Vibrio count and altered
digestive tract providing increased resistance to V. harveyi infection in L.
vannamei after application of feed probiotic Lactobacillus plantarum.
Microbial
Interventions through Biofloc Technology
Biofloc technology
is built on the advance of microbial flocs in suspension by using continuous
vigorous aeration to allow aerobic decomposition (Avnimelech and Weber 1986) as
well as on the principle of flocculation (co-culture of heterotrophic bacteria
and algae) within the system. Such a condition eases the growth of dense
microorganisms that purposes as an in
situ bioreactor and upholds water quality and also exploited by fish as a
protein source (Crab et al., 2009). Further addition of carbohydrate arouses
the growth of heterotrophic bacteria which uptake nitrogen from water and in
turn yields microbial proteins (Avnimelech et al., 1989, 1994 and 1999). Biofloc technology established aquaculture
as equated to conventional aquaculture techniques offers more economical substitute
and sustainable techniques in terms of minimal water exchange and reduced feed
input making it a low-cost sustainable technology for sustainable future
aquaculture development. Heterotrophic bacteria immobilize toxic nitrogen more
rapidly and efficiently as compared to autotrophic counterparts (Hargreaves,
2006) as biofloc are dominated by heterotrophic bacteria hence more efficient
in managing toxic nitrogen in the pond. Biofloc technology based culture system
requires least or no water exchange in aquaculture systems through maintaining
optimum water quality within the culture unit while producing low cost protein
rich bioflocs as supplementary feed for aquatic organisms.
Carbon: nitrogen ratio plays a major
role in biofloc development in establishing a population of heterotrophic
bacteria. Biofloc based aquaculture is a sustainable solution for the
development of aquaculture and this technology is fully based on the concept of
encouraging microbial floc through optimum carbon nitrogen (C/N) ratio (Avnimelech,
1999 and Crab et al., 2007). The control of inorganic nitrogen
accumulation in the pond is based upon the utilization of nitrogen by bacterial
cells by taking energy from the carbon source. The relationship of carbon
source, ammonia immobilization and production of microbial proteins depends on
the microbial conversion coefficient (C/N ratio) and found to be effective in
reducing toxic nitrogen (Ammonia-N and Nitrite- N) in shrimp and tilapia culture
after addition of carbon source (Avnimelech, 1999). A ratio of 10:1 to
20:1(mostly) is used depending upon the type of carbon source and also the nature
of culture practices.
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Diverse
organic carbon sources (glucose, cassava, molasses, wheat, corn, sugar bagasse,
sorghum meal, etc.) are used to boost production and to upsurge the nutrient
dynamics through changed C/N ratio in shrimp culture (Avnimelech, 1999). The
nutritional possessions of bioflocs are largely prejudiced by the types of
carbon sources used to produce the flocs (Crab, 2010; Crab et al.,
2010a). Commonly used carbohydrate sources for carbon supplementation are
starch, wheat flour, wheat bran, molasses, acetate, glycerol and glucose. Dissimilar
organic carbon sources support the growth of specific bacteria, protozoa and
algae, and hence inclined the microbial composition and community organization
of the bioflocs and thereby also their nutritional properties (Crab, 2010). Simple
sugars, such as sucrose, outcome in a quicker ammonia removal, while more intricate
carbohydrates require more time for decomposition into simple sugars, thereby
resulting in slower ammonia removal (Ekasari J., 2014).
Microbial
community in Biofloc
Microbial communities are highly
variable in biofloc and depend on several factors. Now a day specific
probiotics for biofloc are commercially available to boost the population of
only beneficial microbes in floc. Ebeling et al. (2006) investigate the
stoichiometry of autotrophic, photoautotrophic, and heterotrophic elimination
of ammonia-nitrogen in the aquaculture system. The study revealed that two
functional classes of bacteria were mainly responsible for water quality
maintenance in minimal exchange, intensive systems viz., heterotrophic
ammonia-assimilative and chemoautotrophic nitrifying bacteria. Avnimelech
(2007) revealed that the number of bacteria in biofloc ponds can be between 106
and 109 per ml of floc which contains between 10 and 30 mg dry matter making
the pond a biotechnological industry. Ju et al. (2008) disclosed the
effect of inclusion of whole floc or floc fraction to the formulated diet in Litopenaus vannamei. The biofloc
collected from L. vannamei tanks
contained 24.6% phytoplankton (dominated by diatoms like Thalassiosira,
Chaetocerous and Navicula), 3% bacterial biomass (two third was gram-negative
and one third gram-positive), a small number of protozoan communities, 98%
flagellates, 1.5% rotifers, and 0.5% amoeba), and 33.2% detritus, and the
remaining quantity was ash (39.25%). Gutiérrez et al. (2016) revealed
that biofloc established with molasses as a carbon source constitute a diverse
group of microorganisms including 18 heterotrophic bacteria species, 11
opportunistic pathogen and seven probiotic bacteria. With molasses and rice
powder as carbon source, out of 17 bacteria species, 6 pathogens, 9 degradative
and 2 probiotics bacteria were detected. It is also observed that as soon as microbial
flocs progress process starts, the degradative heterotrophic and potential
probiotics displace the opportunistic pathogen bacteria that communally causes
infection in cultured fish and crustaceans such as genus: Aeromonas, Pseudomonas, Vibrio, Enterobacter, Klebsiella among
others, because of a competitive exclusion by heterotrophic and probiotic
bacteria and also secretion of a variety of exoenzymes and polymers that
suppress proliferation of pathogens.
Biofloc
technology (BFT) has been effectively applied in aquaculture especially shrimp
farming due to economical and environmental benefits over a conventional
culture system and due to its capability to reduce the accumulation of toxic metabolites
like ammonia by means of the action of heterotrophic bacterial.
farming
Biofloc
technology based shrimp farming requires a huge investment in terms of aeration
and other maintenance. Periphyton based shrimp farming can be used as an
an ernative of Biofloc technology where microbial aggregate grows over a
substrate. Here carbon source such as molasses is added to the water close to the
substrate which is again placed in close proximity to aerator. It allows the
development of beneficial microbial aggregate on the substrate which in turn
eaten by attracting zooplankton and other small organisms. All these form a
food web with shrimp at the top.
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This whole system acts as an in
situ bioremediation and helps in reducing the level of ammonia in the pond
also. Different types of the substrate can be used but the use of aquamats has
shown better results in terms of periphyton biomass and economic suitability (Kumar
et al., 2019). Such aquamats are also durable and can be disinfected
before use.
The concept of aquamimicry is based on
simulating natural estuarine conditions in the aquaculture pond through
supporting zooplankton bloom as supplemental nutrition to the shrimp and also
maintaining the population of beneficial bacteria for water quality. Here carbon
source is fermented (Romano 2017) by using probiotic strains and then the only
liquid portion is applied in the pond. This approach is certainly different
from Biofloc technology in some aspects such as the amount of carbon
requirement is reduced and does not require strict to maintain a strict C: N
ratio and second not allowing the development of Biofloc by removing
sediments. Such a pond mimics the
natural water condition with plenty of zooplankton and microalga population. It
acts as a synbiotics a combination of both pro and prebiotics. This approach
helps in reducing FCR and minimizing the need for water exchange by maintaining
good water quality through the utilization of toxic components such as ammonia
by bacteria and hence reducing the chance of disease outbreak. Such an environment
provides stress free environment to the cultured animal and also discourages
the proliferation of pathogens.
Rice bran or wheat bran is generally
used as a carbon source that is fermented for 24-48 hours after mixing water in
1:5 to 1:10 ration under vigorous aeration after adding probiotic bacteria
which is mostly Bacillus species. Only the liquid portion is collected using
fine sieve or cloth for application in the pond. This liquid is added at the
rate of 1 ppm in case of extensive culture, whereas 2-4 ppm dose is applied for
intensive culture. The amount can be are adjusted based upon the water
turbidity with optimum around 30-40 cm. During grow out period, supplementary
probiotics should be added each month to assistance uphold water quality and to
stimulate the formation of bio colloids (flocs composed of detritus,
zooplankton, bacteria, etc.). Subsequent 15 days after pond stocking with
shrimp, gradually dragging chains or ropes on the pond bottom (but not over the
central drain) is fortified to diminish the formation of biofilms. For
extensive systems, there is commonly no need for further water quality
management or action. For intensive systems, however, there is a necessity to eliminate
excessive sediments (e.g., through a central drain) to a sedimentation pond two
hours after each feeding. Regardless of the system type, the pH is reportedly
stable throughout. Variation of aquamimicry is also developed where fermented
soyabean is used as a feed instead of pellet feed along with the use of
fermented rice bran (Romano 2017). Such practices require further optimization.
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