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This project has received funding from the European Union‘s Seventh Framework Programme for research,

technological development and demonstration under grant agreement No. 311794.



Dr. Barbara Metzler-ZebeliMr. Andor MolnarMs. Janine Scholz

University of Veterinary Medicine Vienna

Dr. Stefan G. BuzoianuDr. Peadar G. Lawlor

Ms. Ursula McCormack

Moorepark Research Centre,Teagasc, Ireland

Improving feed efficiency by understanding the intestinal bacterial

network in pigs and poultry



Introduction



ECO-FCE overview

• Feb 2013 – Feb 2017

• 17 partners

• 7 WPs

• Overall objectives

• improve food security by optimising the feed efficiency in pigs and broilers without

negatively affecting animal welfare and meat quality

• reduce the ecological footprint of the pig and broiler production systems

• WP 3 objective

• to examine the gut structure, function, microbiota and metagenomics in animals

divergent for feed efficiency



Work Package 3

Intestinal structure &

function

Intestinal health

Intestinal microbiota

Animal performance

Genetics

RFILow

MediumHigh

Health & welfare

Meat quality

Genomics



Determination of feed efficiency

Residual Feed Intake (RFI) = difference between observed and predicted feed intake, with lower RFI values indicating greater energy efficiency

Selection of high and low feed efficient animals - based on Residual Feed Intake

Where a is the intercept and b1 and b2 are partial regression coefficients of feed intake (FI) on BW0.75 and body weight gain (BWG), respectively.

RFI = FI [a + b1 * BW0.75 + b2 * BWG]

Other measures of feed efficiencyFeed efficiency = gain (g) / feed intake (g)Feed conversion ratio (FCR) = feed intake

(g) / gain (g)RG = BWG [a + b1 * BW0.75 + b2 * FI]

RIG = (RG/SD RG) - (RFI/SD RFI)



Feed efficiency in monogastric livestock species

Feed efficiency

Genetics Diet Rearing environment Age

Gut commensal microbiota

Substantial variation in feed efficiency between individual animals. Great variation in gut commensal microbiota between individuals.



Role of the intestinal microbiota

• Benefits to the host• intestinal maturation• inhibition of pathogen growth• nutrient salvaging• detoxification• production of vitamins

• Costs to the host• competition for nutrients• immune activation• production of toxins• opportunistic• toxin reabsorption• mucolytic activity



Effect of host microbiota on host metabolism and hormone secretion

Intestinal microbiota can redirect energy

partitioning to adipose tissue and reduce fatty

acid oxidation.

Bäckhed (2011) Ann Nutr Metab 58(suppl 2):44

Implications for feed use efficiency and carcass

composition in livestock animals?



Effect of gut microbiota composition on body weight

Changes in Lactobacillus and Bifidobacterium species

Obese humans & mice:

Firmicutes Bacteroidetes

Low-calorie dietFirmicutes Bacteroidetes

Actinobacteria Bacteroidetes no difference in Firmicutes

Methanogenic archaea

Meat-producing monogastric livestock species are young, fast growing and lean animals

Are the key players the same as in human obesity models ?

wikipedia.org

Requena et al. (2013) Trends Food Sci Tech 34:44

http://en.wikipedia.org/wiki/File:Fatmouse.jpg

http://en.wikipedia.org/wiki/File:Fatmouse.jpg



Chickens



Diet-related cecal microbiota and performance in male chickens

Torok et al. (2011) AEM 77: 5868

Diet is the most influencing factor affecting feed efficiency.

Caecal microbial communities by diet Caecal microbial communities identified as being from birds with improved performance or poorer performance



Batch to batch variation in caecal microbiota of chickens

Stanley et al. (2013) PloS ONE 8(12): e84290

PCA plot of caecal microbiota. The plot is based on between groups (trials) analysis.

3 different batches of chickens

Very different microbiota profiles across chicken batches

Very different feed use efficiencies across chicken

batches

High variation in caecal microbiota partly due to lack of colonisation of the chickens by maternally derived bacteria

Þ High hygiene levels in modern commercial hatcheries remove natural bacteriaÞ Environmental microbiota from transport boxes, first feed and staff people



Fecal community of high and low feed efficient broiler chickens

Singh et al. (2014) J Appl Genet 55: 145

79

127

Low efficient chickens

Proteobacteria (%)

Firmicutes (%)

Bacteroidetes (%)52

28

18

High efficient chickens



Experimental design:• 2 partner institutions (AFBI & Vetmeduni) performed identical chicken experiments with

3 batches of 50/64 chicks• Similar chicken genetic: Cobb 500FF• Similar maize-soybean meal diets (starter, grower, and finisher diets)• No in-feed antibiotics and any other gut health-related additives• Chickens were individually housed• Best and worst feed efficient chickens were identified using Residual Feed Intake

• On day 42, samples were collected for:• Ileal and caecal digesta for metagenomics and microbial metabolites• Tissue of duodenum, jejunum, ileum, caeca for gut function and structure

Characterisation of differences in gut microbiota and gut function of chickens with good and poor feed efficiency

d1 d7 d14 d28 d42

Weighing Weighing Weighing Weighing

d21 d35

Weighing Weighing

Daily feed intake

Metagenomics(faeces)



Ammoniaemission(faeces)



Residual feed intake of good and poor feed efficient broiler chickens

Great variation in residual feed intake and thus in

feed use efficiency.

-300

-200

-100

0

100

200

300

Good Average Poor

(g)

Residual feed intake

MaleFemale

Feed efficiency, P< 0.001Batch, P>0.1



Microbial metagenome of good and poor feed efficient chickens

Under construction

Shotgun sequencing using MiSeq Technology (Illumina)



Jejunal electrophysiological characteristics of good and poor feed efficient broiler chickens

Good feed efficient females showed lower tissue resistance, higher conductance and short-circuit current indicating a higher ion flux and permeability of the jejunal mucosa

• Gut electrophysiology was performed using Ussing chamber technique. • Tissue originated from the distal jejunum.

Influencing factors: Host genome or gut microbiota ?

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

Good Average Poor

Short-circuit current (Isc; µEq/cm2 x hour)

FemalesFeed efficiency, P=0.076

a

ab

b

0

50

100

150

200

Good Average Poor

Tissue resistance (Ω/cm2)

b

a aFemalesFeed efficiency, P=0.020

0

2

4

6

8

10

12

14

Good Average Poor

Tissue conductance (mS x cm2)

FemalesFeed efficiency, P=0.002

a

b b



Pigs



Literature

• Little data available in pigs

• ↓ Bacteroidetes & ↑ Firmicutes in obese pigs (Pedersen et al., 2013)

• ↑ Firmicutes & ↓ β-Proteobacteria in ERS-fed pigs (Haenen et al., 2013)

• Protein, CHO and lipid metabolic pathways affected by intestinal

microbial profile

• mice (Antunes et al., 2011)

• pigs (Mulder et al., 2009)



P

Screening on feed efficiency in pigs

P

weaning d 42 d 84

F F

d 112

F I C

Teagasc × 3 AFBI

Vetmeduni

46 litters

Common genetics

Common & site-specific boars

Common diets

Common protocols

Pigs divergent

for RFI

F – faecalI – ileal digestaC – caecal digesta P – performance



P PPP

Microbiota profilingd 0 (weaning) d 42 d 84 d 126 d 139

F F F F F I C

F – faecal; I – ileal digesta; C – caecal digesta; P – performance

Compositional analysis16S rRNA gene sequencing

FunctionalityShotgun metagenomicsIllumina



Progress on microbiota profiling

• Samples collected

• DNA extracted

• 16S rRNA gene sequencing – results being analysed

• Shotgun metagenomics

• samples being prepared

• results ~ Oct 2014



Manipulation of GIT microbial profileMicrob

ial inocula

tion

Nutrition

Management

Low RFI

Additives



• Anaerobically processed• diluted 1:6• strained• centrifuged (6000 × G for 15 minutes)• frozen at -80°C in 10% glycerol

Inoculation with faecal inoculum from good feed converters

No inoculum

No inoculum

Single inoculation

Multiple inoculation

Inoculum

No inoculum

Single inoculation

Multiple inoculation

Sows

Offspring



• Optimum strategy – inoculum

• Prebiotics – alone or in combination

• Monitoring and sampling of offspring through their lifetime

• performance

• health

• intestinal microbiota

Nutritional intervention



Acknowledgements

• ECO-FCE has received funding from the European Union’s

Seventh Framework Programme for research, technological

development and demonstration (FP7 2007/2013) under grant

agreement No. 311794

• Teagasc Walsh Fellowship Programme



Thank you



ECO-FCE Gut structure, function, microbiota and metagenomics

Objectives:

1. To enhance our understanding of the interactions between gut microbiome and host genome in pigs and chickens. This task will be achieved by employing cutting-edge 16S rRNA-specific and shotgun metagenomics.

2. Using this improved understanding, strategies to improve feed conversion efficiency through gut microbiome manipulation in embryonic and subsequent developmental stages will be developed.

Hypothesis:It is assumed that the gut microbiome of pigs and broiler chickens with good and poor feed use efficiency differs in key members, thereby influencing the intestinal and metabolic host response, production efficiency and host health.



NF-kB

Interactions between gut microbiome and host physiology and health

Bäckhed (2011) Ann Nutr Metab 58(suppl 2): 44; Twarziok et al. (2014) Mol Inf 33: 171

http://edoc.hu-berlin.de/dissertationen/buettner-mirjam-r-2005-01-24/HTML/buettner_html_32688095.gif











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