Phytogenics in poultry nutrition; an important source of antimicrobial, anticoccidial, antioxidant and antiinflammatory compounds
Lecturer Dr. Ilias GIANNENAS Laboratory of Nutrition, Faculty of Veterinary Medicine,
Aristotle University of Thessaloniki, Greece
Future Dynamics in sustainable Animal Nutrition, November 6 & 7, 2014, Hart van Holland conference centre, Nijkerk, The Netherlands
SCHOTHORST FEED RESEARCH –
80 YEAR ANNIVERSARY CONFERENCE
Presentation outline  Introduction
 Phytogenics
 Plant bioactive compounds and health implications





Antimicrobial effect
Antiprotoozoal effects
Antioxidant effect
Antiinaflammatory effect
Other effects
 Conclusions
 Perspectives for future research
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
1
Demands
Consumer
Producer
Good quality food
No residues
Optimum performance
Healthy crops
Antibiotic Growth Promoters
& Coccidiostats
Natural Alternatives
Growth promoters as feed additives
 The use of feed additives as a mechanism for manipulating growth
promotion in farm animals has been extensively investigated since 1950s
(Beeson and Perry, 1952)
Resistance to antimicrobials !
 Recently, animal production in many countries has faced restrictions in
the use of antibiotic growth promoters
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
2
Consequences of AGPs ban on gut health
Dysbacteriosis / Wet Litter
problems
Decreased digestion
Reduced weight gain
Impaired feed conversion ratio
Ban on AGPs
2006
Necrotic enteritis
Coccidiosis
Intestinal necrosis
Intestinal necrosis
Mortality
Estimated cost
= 0.05$ / bird
Mortality
Alternatives for bacterial diseases
•Necrotic enteritis ‐ significant losses, cost, especially after the AGP ban
•Enteritis (E. coli infection, Salmonellosis) – mortality, bacteria transfer •Omphalitis (Navel infection) – mortality, depressed growth rate
•Chronic respiratory disease – mortality depressed growth rate
•Avian Campylobacter infection – bacteria tranfer, endangered food chain •Foot pad disease – depressed growth rate
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
3
Alternatives for protozoal diseases ‐ Coccidiosis
 Coccidiosis is a significant endemic disease in poultry
3
Billions
 The financial loss to
the poultry industry
worldwide circa
3 billion USD/year
2008
 Cost mainly due to prophylactic or therapeutic in‐feed medications and also
as a result of the disease impact on birds’ health
(Williams, 1999; Bozkurt et al, 2013)
Thymol
Hippocrates, the ‘father of medicine’, used plant extracts and prescribed perfume fumigations
Phytogenics
Thyme
Piperin
e
Black Pepper
“Products” of plant origin
o Botanicals
o Plant extracts and essential oils
o Pure compounds
 Last decades several in vivo studies have been carried out to evaluate the effectiveness of essential oils to manipulate growth performance and improve nutrient utilization on poultry (Brenes and Roura, 2011)
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
4
Phytogenics ‐ Activities & substances
 Antimicrobial
 Bactericide
 Parasiticide
 Fungicide
 Insecticide
 Anti‐oxidant
Bioactive substances
 Carvacrol
 Thymol
 Eugenol
 Limonen
 Vanilin
 Cinnamaldehyde
 Anti‐inflammatory  Tannins
 Feed intake increase
 Stimulating digestive secretions
Antimicrobial mechanism of EOs
•The hydrophobicity of EOs enables them to partition in the lipids of the cell membrane and mitochondria, rendering them permeable and leading to leakage of cell contents
•Disintegrate the outer membrane of Gram‐
negative bacteria releasing lipopolysaccharides (Burt, 2004)
Synergistic effect provides an increased intensity caused by the combination of two or more substances on an organism
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
5
Bacteriostatic/Bactericidal effects of EOs
Rank
Chemical group
Component
Essential Oil
1
Phenols
Carvacrol, thymol, eugenol, gaiacol
2
Aldehyde
cinnamaldehyde
Inner bark of cinnamon tree
3
Monoterpénol C10
1-terpinène-4-ol
Tea tree
4
Aldehydes
Geranial, citronellal
Lemon grass
5
Cetones
Verbenone, menthone, carvone
Rosemary, aniseed, mint
6
Ether
Estragol, anethol
Basil, aniseed, star anise
7
Oxydes
Eucalyptol
Eucalyptus
8
Terpenes
Pinenes
Pines, firs
Oregano, thyme, clove, cinnamon leaves, …
Knobloch et al., 1989; Franchomme et al., 1990; Dogna, 1990; Inouye et al., 2001; Friedman et al., 2002;
Hernander-Ochoa, 2005
In vivo studies
 On this basis, it is logical to consider essential oils application in vivo as the intake of essential oils could replace antiobiotics
 Vogt, early as 1981, used hers and spices in poultry nutrition
 Essential oils, preparations of herbal extracts & blends with pure phenolic substances showed a reduction of colony forming units of many bacteria such as Clostridium spp
 Herbal feed supplements can affect gastrointestinal microflora composition and population. Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
6
Phytogenics as growth promoters
 Dietary use of:
 Oregano plant
 Oregano extracts/ essential oils
 Carvacrol, thymol
 Initial research yielded: 82 papers
 Suitable for this review: 47 papers
Parameter
N of
manuscripts
Maximum
deficit %
Maximum
benefit %
BW
9
-6.74
+23.00
BWG
8
N.S.
+14.11
Animal type
No of
manuscripts
Chicken broilers
26
Chicken hens
2
Turkeys
3
Quail
2
Rabbits
1
Pigs
9
Sows
1
6
-15.07
+9.20
Cows
1
13
-5.45
+17.19
Ewes
1
Mortality
0
N.S.
N.S.
Lambs
1
Egg
0
N.S.
N.S.
Milk
1
N.S.
+35.39
FI
FCR
Giannenas et al., 2013
Anti ‐parasitic, ‐protozoal & ‐fungal mechanisms of EOs
 Phenols exhibit oocysticide activity as disinfectants in vivo & in vitro
tests (Williams, 1997)  Phenols are targeting against cell membranes affecting cell wall structure, internal cell membranes and enzyme activities
 Polyphenols (tannins) have direct effects to parasites, but also to the eggs
 Active substances react with sulfhydryl groups, which are indispensable for the fungal growth
 Formate of charge transfer complexes with electron donors in the fungus cell that  lead to inhibition of cell division and  interference with cell metabolism
 inhibition of the fungal‐cell‐wall synthesizing enzymes Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
7
Current interventions to control coccidiosis in poultry
 Chemotherapeutics (antibiotics ‐ chemicals)
 Control of coccidia has been greatly dependent on the use of chemotherapeutics agents
 It has proven success in many parts of the world, due to its ease of use and the ability to provide uniform treatment and prevention, however
On some occasions, might be toxic to the birds
The constant pressure to reduce the dependence on antimicrobials including the anticoccidial drugs, because of  High cost  Public health concern for drug residues in poultry meat, eggs, foods
 Resistance development to drugs by Eimeria species
Resistance to anticoccidial drugs worldwide
• Approved drugs= Ionophores, synthetic chemicals, sulfonamides, mixed
products
• Resistance ”the ability of a parasite strain to survive and/or multiply despite the administration and absorption of a drug in doses equal to or higher than those usually recommended”
• Development of resistance to all anticoccidial drugs
• Widespread occurrence in US, EU, South America, South Africa, China (1974‐2004)
• Cross resistance to new substances with similar mode of action!
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
8
Alternative control of coccidiosis in poultry; Phytogenics?
Plants/extracts/compounds
Scientific name
Challenge/beneficial effects/mode of action
Literature
Oregano aerial parts and
essential oil (containing
carvacrol and thymol)
Origanum
vulgare ssp.
hirtum
Eimeria mix (8 species): lesions, oocyst
shedding, FCR; E. tenella: body weight gain,
FCR, lesions, oocyst shedding
Batungbacal et al.
(2007), Giannenas et
al. (2003,2004)
Quinghao dried herb;
artemisinin; 1,8-cineol;
camphor; A. sieberi
petroleum ether extract of
aerial parts; A. afra
acetone/water extract from
aerial parts
Green tea leaves
China bark tree extract,
Quinine
Turmeric spice rhizome,
Curcumin
Artemisia annua, E. tenella: lesions, oocyst shedding; E.
A. sieberi., A.
acervulina: oocyst shedding (not E. maxima);
afra
Eimeria mix (E. tenella, E. maxima, E.
acervulina): FCR; induction of oxidative
stress.1,8-cineol and camphor: body weight
gain, lesions
Camellia sinensis
Cinchona
succirubra
Curcuma longa
E. maxima: oocyst shedding
E. tenella, E. meleagrimitis: sporozoite invasion
in vitro
E. maxima: lesions, body weight gain; E.
acervulina (not E. tenella); antioxidative
Allen et al.
(1997,1998), Allen &
Fetterer (2002), Arab
et al. (2006), Naidoo et
al. (2008)
Jang et al. (2007)
Christaki et al. (2004),
Fayer (1971)
Allen et al. (1998),
Allen & Fetterer(2002)
(1/2)
Bozkurt et al (2013)
Anticoccidial phytogenics
Plants/extracts/compounds
Challenge/beneficial effects/mode of action
Literature
Oriental plum; Japanese plum Prunus salicina
Scientific name
E. acervulina: body weight gain, oocyst
shedding, IFN-γ and IL-15 (mRNAs) of
intraepithelial lymphocytes, spleen cell
proliferation; phenolics and antioxidants
Lee et al. (2007)
Sugar cane extract
E. tenella: body weight gain, oocyst shedding,
lesions, antibody response
E. tenella: body weight gain, diarrhoea,
mortality, lesions, oocysts shedding
E. tenella: weight gain, mortality, bloody
diarrhoea
Eimeria mix (E. tenella, E. maxima, E.
acervulina): FCR, oocyst shedding
El-Abasy et al.
(2003)
Florou-Paneri et al.,
2004
Youn & Noh (2001)
E. tenella: body weight gain, mortality, lesions;
Eimeria mix (E. tenella, E. maxima, E.
acervulina): FCR
Naidoo et al. (2008),
Wang et al. (2008)
Olympus tea
Saccharum
officinarum
Sideritis scardica
Ku Shen root decoction
Sophora flavescens
Wild garlic acetone/water
extract from whole plant;
marasmine
Tulbaghia violacea
Grape seed proanthocyanidin
extract; ethanol/water extract
from pomace
Vitis vinifera
Artemisia, Thyme
(2/2)
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
Naidoo et al. (2008)
Bozkurt et al. (2013)
9
E. tenella in vitro ::: screening
a
b
b
c
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
10
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
11
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
12
b
c
c
d
a
E. tenella in vivo ::: challenge trial
Group
Code
Description
1
UU
untreated, uninfected
2
UI
untreated, infected
3
Las
4
T
5
Tenc
Inclusion
rate
(g/ton feed)
Lasalocid
75
Thymol
300
Thymol, matrix
encapsulated
140
 2E+04 E. tenella
oocysts per bird at day
14 (crop intubation)
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
13
E. tenella in vivo ::: challenge trial
UU
UI
Las
T
Tenc
BW (g)
Day 35
1865a
1780bc
1842ab
1710c
1831ab
FI (g)
2986a
3206b
2931ac
2737 c
3046ab
FCR (FI/BWG)
1,641a
1,849c
1,632a
1,644a
1,706b
c
b
a
a
a
ab
FCR (FI/BWG)
BW (g)
a
ab
bc
c
UU
UI
Las
T
Tenc
E. tenella in vivo ::: challenge trial
UU
UI
Las
T
Tenc
LS d21 (0-4)
0,00a
2,67b
1,75b
2,38b
2,00b
LS d35 (0-4)
0,17a
2,17b
0,67a
2,00b
1,67b
0
3
2
2+
3
0,0ac
86,4b
19,4c
31,1b
4
2
2
2
DS d17-21
OPG reduction (%)
Mortality (birds/30)
0
Normal villae
Cocci – affected villae
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
14
Effects of coccidia on intestinal villi
Normal
Cocci affected
They need 96 hours to repair!
Do chickens have this time?
Anti‐oxidant mechanisms of EOs
 Phenolics react with free radicals and reactive oxygen species like tocopherols
 Detoxify and protect cell and organs
 Spare effect on tocopherols like ascorbic acid
 Prooxidant activity at certain levels like ascorbic acid?
 Antioxidant mechanisms not yet fully understood
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
15
Results on antioxidant activity 1600
CONTROL
OR5
OR10
OR5-TΟC
OR10-ΤΟC
ΤΟC
MDA (ng/g)
1200
800
400
0
0
90
180
270
Incuba tion tim e (m in)
MDA levels in breast tissue submitted
to iron-induced lipid oxidation
Anti‐inflammatory effects of EOs
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
16
Oregano substances
downregulated ileal IL‐6 and tonsil LITAF, IFN‐γ, TLR4 and IL‐10 gene expression
& exerted a significant anti‐inflammatory effect
E. tenella in vivo ::: screening
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
17
Phytogenics – Digestibility enhancer on chicken
 Thymol, cinnamaldehyde or a commercial preparation of essential oil components were shown to increase amylase activity in the intestinal digesta of female broilers (Lee et al., 2003).
 Increase lipase activity in the pancreas and intestinal wall with a plant extract consisting of capsaicin, cinnamaldehyde and carvacrol with FCR
improvement in broilers (Jamroz et al.,2005).
 Dietary piperine (pungent principle of black pepper) stimulated the digestive enzymes of pancreas and enhanced the digestive capacity and reduced the gastrointestinal feed transit time (Srinivasan, 2007).
 Plant extracts from a mixture of oregano, cinnamon and pepper or from sage, thyme and rosemary improved apparent whole‐tract and ileal
digestibility of nutrients in broilers (Hernandez et al., 2004).
Adverse effects of aromatic plants
 In a work with oregano ground plant, although diet supplementation at 5.0 and 7.5 g/kg of feed protected birds
 Oregano at 10 g/kg, had non‐protective role possibly due to action of its phenolic constituents that in high doses might have toxic effects (Giannenas et al., 2004)
 Phenolic compounds may exert their activity on the host enterocytes, too (Weber and De Bont, 1996)
 Carvacrol and thymol might have breaking effect on the upper layer of mature enterocytes of mucosa, which are already affected by intracellular pathogens
 Hydrophobic character of carvacrol suggests interaction with membranes. When concentration of carvacrol increases, it is expected to accumulate and interact in the phospholipid belayer, affecting membrane fluidity Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
18
Explanations for inconsistent results  Evidence on biological active properties of plants is not consistent with the expectations arising from tradition  Majority of ethno pharmacology reports originate from ruminants, as the main livestock species that generate income in poor countries
 When such plants are tested in monogastrics, part of the reported variation may be due to physiological difference between ruminant and nonruminant
animals (Githiori et al., 2003)
 Essential oils due to their potent nature should be used as low as possible levels in animal nutrition
 They may lead to feed intake reduction or gastrointestinal microflora
disturbance or toxicity Explanations for inconsistent results (2)
 Variation observed in the experimental protocols and level of supplementations
 Various effects on gut microflora and intestinal protozoa but also on animal metabolism
 Variable composition. For example, concentration of two predominant
components of thyme essential oils, i.e. thymol and carvacrol have been
reported to range from 3% to 60% of total oils
 Observed lack of growth‐promoting effect in some trials may relate to composition of basal diet and/or environmental conditions
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
19
Safety Issues ?
 Food grade listed as GRAS (Generally Recognized As Safe)
 Appear on EU Register of Feed Additives
(Reg.1831/2003)
 Residues have to be monitored
Share of phytogenics in EU market
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
20
Production of phytogenics
Authentification of raw materials
Chemical standardization
Process chemistry: scale up
In vitro
studies
In vivo
studies
Field Trials
Quality
control
GMP production and marketing
Future perspectives
New investigations are warranted on the following topics:
• mechanisms of action for promising compounds in combinations (e.g. EOs with health benefits, +probiotics, +other compounds e.g. organic acids) • bioavailability and pharmaco‐kinetics of EOs + other feed additives e.g. enzymes • zootechnical and biological effects of combinations of alternatives based on the fact that GIT disorders post‐hatching display highly complex aetiology and mechanisms, interaction with immunity
• negative interactions between feed components and added alternatives OR among alternatives must be understood
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
21
Novel Search
 Numerous native herbs are rich in phenolic compounds
 However, multidisciplinary research is necessary with the interactions of research Institutes and, the Feed Additive industry, the Feed industry and the Poultry industry in order novel substances to be tested!
 Progress can be achieved by considering the various aspects of medicinal plants together and thus exploit their complementarity
 Emphasis and additional ongoing research for
 performance,  immunity  behavioural observations
Main References
 Aromatic plants as a source of bioactive compounds: E. Christaki, E. Bonos, I. Giannenas, P. Florou‐Paneri, Agriculture, 2012
 An update on approaches to control coccidia in poultry using botanical extracts: M.
Bozkurt, I. Giannenas, K. Kucukyilmaz, E. Christaki, P. Florou‐Paneri, British Poultry
Science, 2013
 Essential oils and their applications in animal nutrition ‐ Invited Review: I. Giannenas,
E. Bonos, E. Christaki, P. Florou‐Paneri, Medicinal and Aromatic plants, 2013
 Oregano supplementation in farm animal diets: effects on productive parameter: Ι.
Giannenas, Ε. Bonos, E. Christaki, P. Florou‐Paneri. 17th Congress of the European
Society of Veterinary and Comparative Nutrition, Ghent, Belgium, 2013.
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
22
Thank you for your attention !
Thessaloniki, Greece
Future Dynamics in sustainable Animal Nutrition, Nov 7, 2014
23