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Immunomodulatry and growth responses of Nile Tilapia fingerlings to dietary extract of Moringa flower
Esayas Welday Tekle1,2 & N. P. Sahu1
1 Fish Nutrition, Biochemistry and Physiology Division, Central Institute ofFisheries Education, Panch Marg, Off Yari Road, 400061 - Mumbai, India
2 College of Marine Sciences and Technology, Massawa, EritreaE-mail: [email protected]
Aquaculture Europe 2015European Aquaculture Society20 -23 October, 2015, Rotterdam, Netherlands
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Outline
Bottomline Aquaculture Aquafeeds
Challenges Alternatives Dietary Moringa extract Effect on
Growth Enzyme activities Immunological
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Tilapia Aquaculture
Aquaculture
Tilapia aquaculture Most widely distributed
(>135 countries) 2nd in production
Species of choice Hardy Low trophic level Flesh quality (fillet or
whole fried)
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Farming and Production of O. niloticus (FAO, 2010)
Asia
Africa
Americas Others
Aquaculture production of tilapia by countryin million tonnes 1950–2009 (FAO, 2012)
Aquaculture production of tilapia by speciesin million tonnes as reported by the FAO, 1950–2009[1]
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Challenges to industry sustainability
Intensification of farming systemsNutrient loadWater quality – stress
Feed and feed relatedEconomicEnvironmentalHuman health
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ROS (FR)Oxidative stressInfectious diseases
Pathogens (opportunistic)
Stressful environment
Host (fish)
Overcrowding
Husbandry
Physicochemical (Poor WQ)
Parasite infestation
?
Modified from:Source: http://www.thefishsite.com/articles/1570/an-introduction-to-fish-health-management/ accessed on 11/10/2015.
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Synthetic antioxidants BHT BHA Ethoxiquine
Chemoteraupitics Various antibiotics Disinfectants
Risks Bioacumulation Residues Development of resistant microbial strains
Environmental and human health concern
Restrictions
Cont. …
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Alternatives
Plant/herbal extracts Natural Minimal or no negative impact
Antioxidant properties Mitigate oxidative stress – scavenge ROS Prevent lipid peroxidation - rancidity
Antimicrobial effect Enhance immune system Combat microbes
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Moringa
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Moringa?
Deciduous plant species Moringa spp. (Moringacea
family) Moringa oleifera – most
widely distributed worldwide “drumstick tree” “horse radish tree” “ben oil tree”
Native to Himalayan region of India Distributed worldwide
(resistant to drought)
Source: http://miracletrees.org/moringa_powder_content.html
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Medicinal use of Moringa
Ayurveda (more than 300 diseases) The roots, leaves, gum, flowers, barks and
seeds/pods have active chemical constituents:
Nitrile mustard oil glycosides thiocarbamate glycosides (Anwar et al., 2007).
• Diuretic lowering• cholesterol lowering• Antiulcer• Hepatoprotective• cardiovascular
protective property
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Cont. …
Other Phytochemicals with powerful antioxidant & antimicrobial attributes pterygospermin Kaempferol Quercetin Rutin caffeoylquinic acids antioxidant vitamins - C, E, and A carotenoids - lutein, alpha-carotene and beta-
carotene, xanthins, and chlorophyll
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Objectives
Immunomodulatory
Growth performance and nutrient utilization
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1. Extraction procedure
CentrifugingConcentrating Extract (lyophilized)
Pulverized Filtered Shaking (24 hrs) Filtering
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2. Experimental design and methodology
60-days growth period 4 diets (isonitrogenous, isocaloric) basal diet (control) – without extract Three ext. inclusion levels (0.25%, o.5% and 1%)
Diet Denotations Extract level (%)
C (control) 0
EF.25 0.25
EF.5 0.5
EF1 1
EF = ethanolic flower exract (70% ethanol)
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Cont. …
Experimental fish
60-day growth study 120 fish (8.78±1.36g ) in 12 plastic tanks (80 L)
Challenge experiment
Fish were exposed to A. hydrophila for 10 days after the 60-days growth period
20 fish per treatment in duplicate
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Cont. …
After 60-days growth period and 10-days challenge:
Growth parameters
Metabolic and antioxidative stress enzymes
Immunological parameters
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Results and discussion
Diets
Growth parameters Control EF.25 EF.5 EF1
Weight gain 44.90±0.48a 75.90±1.60c 68.29±1.23b 45.92±0.38a
FCR 2.13±0.02c 1.32±0.03a 1.42±0.02a 1.68±0.09b
SGR 0.68±0.03a 1.06±0.03c 0.92±0.02b 0.67±0.05a
Survival (%) 60.00±0.00a 90.00±0.00c 76.67±5.77bc 73.33±5.77ab
PER (%) 1.58±0.03a 2.53±0.05c 2.35±0.03c 1.99±0.10b
Energy retention (%) 12.93±1.08a 24.06±0.58d 20.53±0.18c 17.57±0.34b
Protein retention (%) 24.74±0.33a 51.69±1.22g 44.19±0.03e 43.68±0.52e
Survival(postchallenge) 29.63±6.41a 59.26±6.41c 51.85±6.42b 33.33±11.11a
Table 2 Growth and nutrient utilization
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Some pictures taken after challenge with A. hydrophila
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2. Metabolic enzymes activitiesMDH & LDH
d
c
ab
c
ab
c
a a
b
b
c
c
dd
LDH – anaerobic energy production; conversion of pyruvate (glycolysis) to lactateMDH – energy production from non-carbohydrate sources (TCA cycle); retarded growth
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Transaminases
a
ab
c c
b
c
d
aa
ab b
b
c
d
ALT AST
ALT & AST • Synthesis and deamination of AA.; gluconeogenesis• Higher levels - energy utilization from non-carbohydrate sources – slow down increase in
body mass
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Antioxidative stress enzymes
Treatments
Liver GillPre-
challengePost-
challengePre-
challengePost-
challengeControl 30.41±0.96dA 34.36±1.53cB 28.20±1.70dA 32.03±0.26dB
EF.25 7.01±0.37aA 10.86±1.01aB 15.95±0.80a 13.10±0.74a
EF.5 20.52±0.33c 21.03±0.56b 22.72±1.79bA 26.45±0.71bB
EF1 15.82±1.80b 20.79±0.95b 25.83±1.39c 28.95±0.15c
SOD (U/min/mg protein)
Control 1.94±0.06aB 1.31±0.03aA 1.45±0.1aA 2.12±0.01aB
EF.25 5.28±0.19dB 2.47±0.46cA 2.86±0.3dA 4.21±0.08cB
EF.5 4.44±0.19cB 2.05±0.10cA 1.92±0.2bA 2.92±0.02bB
EF1 3.57±0.15bB 2.00±0.05bA 2.34±0.14cA 2.24±0.00aA
CAT (mmols/min/mg protein)
SOD – cleaving superoxide radicals into elemental O2 and H2O2CAT – decomposes H2O2 into O2 and H2O
Lowercase superscripts in a column; uppercases in a row = significant difference (P<0.05)
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Cont. …
a
abb
c
a
a
cbc
b
c
cba
c
a
GST – catalyzes conjugation of glutathione to toxic cpds. (epoxides, aliphatic, aromatic and heterocyclic radicals, etc.) – measured as µM of CDNB released
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3. Immunological
Treatme
nts
NBT Lysozyme activity
Prechallenge Postchallenge Prechallenge Postchallenge
Control 0.721±0.013aB 0.604±0.009aA 0.367±0.025aA 0.490±0.030aB
EF.25 1.526±0.018cB 0.877±0.040cA 1.983±0.0.085dA 2.243±0.045cB
EF.5 1.093±0.025bB 0.684±0.031bA 1.080±0.076cA 1.533±0.015bB
EF1 0.759±0.013aB 0.677±0.033bA 0.790±0.060bA 1.450±0.120bB
NBT = Nitroblue tetrazolium (absorbance 620nm) – respiratory burst activityLysozyme activity = A450 – a measure of bacterial lysis by blood cells.
Table 4
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Cont. …
Treatments
Phagocytic activity (%) Antiprotease activity (%)
Prechallenge Postchallenge Prechallenge Postchallenge
Control 46.170±1.830aB 34.627±1.375aA 85.633±0.575aB 77.070±0.520aA
EF.25 94.547±.815dB 70.000±2.082dA 94.253±0.385cB 89.540±0.360cA
EF.5 86.670±0.000cB 63.750±2.165cA 90.997±0.575bB 81.897±0.515bA
EF1 78.887±0.964bB 59.167±0.722bA 84.673±0.765aB 76.210±0.690aA
Table 5 Serum phagocytic and antiprotease activities (%) of the various experimental groups
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Conclusion
Growth parameters, enzyme activities and immunlologicalanalysis revealed the EF.25 found to be most effective Recommended as antioxidant and immunostimulant Replace synthetic antioxidants and drugs
As the extract inclusion level increases, performance decreased In some instances less than the control May be due to the toxic effect of the extracts at higher doses Cytotoxicity, histopathological study should be carried out
Mechanism of action, at molecular level
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References Anwar F., Ashraf M. & Bhanger M.I. (2005) Inter-provenance variation in the composition
of Moringa oleifera oilseeds from Pakistan. Journal of the American Oil Chemists' Society82(1), 45-51.
Asaolu, V. O. & Okewoye, A. T. (2013) Moringa multinutrient block supplementation effects on feed Utilization by West African dwarf goats fed a basal diet of Cassava peels. Science Focus 18 (1), 63-72.
Bauer A.K., Dwyer-Nield L.D., Hankin J.A., Murphy R.C. & Malkinson A.M. (2001) The lung tumor promoter, butylated hydroxytoluene (BHT), causes chronic inflammation in promotion-sensitive BALB/cByJ mice but not in promotion-resistant CXB4 mice. Toxicology169(1), 1-15.
Fawole F.J., Sahu N.P., Pal A.K. & Ravindran A. (2015) Haemato-immunological response of Labeo rohita (Hamilton) fingerlings fed leaf extracts and challenged by Aeromonas hydrophila. Aquaculture Research June 25 DIO:10.1111/are.12829.
Moringa oleifera extract against waterborne Pb stress (Sirimongkolvorakul et al., 2012) Siddhuraju P. & Becker K. (2003) Antioxidant properties of various solvent extracts of
total phenolic constituents from three different agroclimatic origins of drumstick tree (Moringa oleifera Lam.) leaves. Journal of agricultural and food chemistry 51(8), 2144-2155.
Sirimongkolvorakul et al., 2012Moringa oleifera extract against waterborne Pb stress Tekle E.W., Sahu N.P. & Makesh M. (2015) Antioxidative and antimicrobial activities of
different solvent extracts of Moringa oleifera: An in vitro evaluation. International Journal of Scientific and Research Publications 5(5), 255-266.
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