For Internal Use Only

The science

behind

peptidoglycans

Pietro Celi (DVM, PhD)

Global Science Lead Eubiotics

DSM Nutritional Products, Animal Nutrition and Health

For Internal Use Only

PRESENTATION OUTLINE

Gastrointestinal functionality

The problem Peptidoglycans

Can we fix it?Muramidase and gastrointestinal

functionality

Take home messages

GASTROINTESTINAL FUNCTIONALITY#FEED4GAIN

For Internal Use Only

#FEED4GAIN

GASTRO-INTESTINAL (GAIN) FUNCTIONALITY

‘a steady state where the

microbiome and the intestinal

tract (host) exist in symbiotic

equilibrium and where the

welfare and performance of the

animal is not constrained by

intestinal dysfunction’

Welfare

Diet

GIT

microbiota

Digestion &

absorption

GIT mucosa

Immune

status

Gastrointestinal

Functionality

For Internal Use Only

Animal performance

Antimicrobial resistance

(#AMR)

#Welfare & Environment

#Food Safety

#sustainableanimal

production

WHY GASTROINTESTINAL FUNCTIONALITY?

Gastrointestinal Functionality is the single most important factor

impacting poultry producers profitability and license to produce

THE PROBLEMWASTE IN THE GUT

For Internal Use Only

Intensification

Stocking density

Removal of AGPs

Incidence of intestinal problems

CHALLENGES IN THE POULTRY INDUSTRY

For Internal Use Only

INTESTINAL CHALLENGES

GASTROINTESTINAL FUNCTIONALITY

Necrotic Enteritis & coccidiosis

Salmonella & Campylobacter

Malabsorption

Bone mineralization

(skeletal health)

For Internal Use Only

GASTROINTESTINAL FUNCTIONALITY

Welfare

Maximise value from

feed

Robust immune system

Nutrient digestibility

Management practices

Is there something new out there?

WHAT IS NEEDED

THE GREATEST THREAT IS THE ONE THAT GOES UNOBSERVEDPEPTIDOGLYCANS

For Internal Use Only

GIT MICROBIOTA: DEAD OR ALIVE?

UNLOCKING THE HIDDEN POTENTIAL

Gadde, U., Lillehoj, H. 2017. Antibiotic Alternatives- A Review. Animal Health Research Reviews.

Millions of microbes are swallowed continuously

Billions of microbes are present in the GIT tract

Microbes are continuously dying off

What happens to the dead bacteria?

Completely overlooked area= bacterial cell debris

For Internal Use Only

ALL PARTS INTERACT WITH THE HOST CELLS IN THE GIT

A HOLISTIC VIEW OF MICROBIOTA INCLUDES DEAD BACTERIA AND CELL DEBRIS

Dead bacteria

(whole cells + fragments)

Live bacteria +

For Internal Use Only

PEPTIDOGLYCANS ARE MAJOR BACTERIAL CELL WALL COMPONENTS

Gram-bacterium

Gram+bacterium

For Internal Use Only

When bacteria and their cell debris accumulate on the intestinal mucosa

• impair absorption

• enterocyte damage

• increased intestinal permeability

PEPTIDOGLYCANS IN THE GUT HINDER GASTROINTESTINAL FUNCTIONALITY

For Internal Use Only

GUT MICROBIOTA LOAD

increase in the GIT microbiota

load

increase in bacterial cell

debris

more PGNs

Source: Maresca, 2013

Dietary protein content

Starch content

NSP levels

Antinutritional factors

Micotoxins

Environmental stress

Pathogen load

MORE BUGS, MORE PEPTIDOGLYCANS

For Internal Use Only

GIT microbiota is composed of both gram- positive (G+)

and gram-negative (G-) bacteria.

When cell walls accumulate in the GIT tract they

become “intestinal waste”

Nutrient digestion and absorption are particularly

prone to the adverse impact this of this cell

debris.

SUMMARY

GASTROINTESTINAL FUNCTIONALITY

Bacterial Overgrowth Impacts Nutrient Absorption

CAN WE FIX IT?YES WE CAN

For Internal Use Only

DISCOVERED AND DEVELOPED BY THE DSM/NZ ALLIANCE

THE DISCOVERY OF MURAMIDASE 007

Expression

Diversity

selection

In silico

In vivo

FCR

Purification Production feasibility

Stability tests

Counter screen

Antimicrobial

potency

Activity screen

Peptidoglycan hydrolysis

Idea review

Project kick- off

Upscale and formulation

ProductLead

enzyme

Iteration cycles

Muramidase 007 was selected as the best out of >

1´000 muramidase candidates

For Internal Use Only

NOT ALL MURAMIDASES ARE THE SAME

THE SOLUTION TO THE PROBLEM

Muramidase 007 specifically catalyze the hydrolysis

of peptidoglycans

MURAMIDASE 007HEWL

For Internal Use Only

NOVEL MURAMIDASE

THE SOLUTION TO THE PROBLEM

Activity on cell debris peptidoglycan component

No antimicrobial potency

Optimal pH activity range 4-7.5

Stable under gastric conditions

Heat stable

For Internal Use Only

BEFORE AND AFTER MURAMIDASE INCUBATION

PEPTIDOGLYCANS

For Internal Use Only

Removing bacterial cell debris promotes optimal digestion and absorption

Releasing amino acids and sugars for host and microbiota

Less damage to the intestinal mucosa

• less energy for tissue repair

• more energy for production purposes

HYDROLYZING PEPTIDOGLYCANS MAY RELEASE A HIDDEN POTENTIAL

UNLOCKING THE HIDDEN POTENTIAL

A hidden potential inside the gastrointestinal

tract = Bacterial Cell Debris

MURAMIDASEIMPROVES GASTROINTESTINAL FUNCTIONALITY#FEED4GAIN

For Internal Use Only

MURAMIDASE INCREASES ABSORPTION CAPACITY

Male Ross 308 fed with diets based on SBM/ Corn/ Rice bran (5%)/ MBM + HiPhos + Carophyll Yellow Challenge: 15x RD Bio-Coccivet R®

*AGP = Enramycin (10ppm Starter and Grower + 5ppm Finisher)

a b ab b

L. Cardoso, 2016

Measurement with iCheck™ Carotene (quick test).

Bittencourt et al., 2018

For Internal Use Only

FITC-Dextran

Lower FITC in blood =

better gut permeability

Cardoso, 2017

FITC-DEXTRAN AS A MARKER FOR GUT PERMEABILITY: NUTRIENT

ABSORPTION AND BACTERIA TRANSLOCATION

MURAMIDASE AND NUTRIENT ABSORPTION

For Internal Use Only

Source: BR 170330 (UFPR) - A. Maiorka, et al., 2017. Unpublished data

MURAMIDASE IMPROVES INTESTINAL PERMEABILITY AND ABSORPTION CAPACITY

For Internal Use Only

Source: ME-01/17VN Aureli & Perez Calvo. Unpublished data

INTESTINAL HISTOMORPHOLOGY: VILLI LENGTH AND CRYPT DEPTH, AND

MUCOSA THICKNESS

MURAMIDASE DECREASES MUCOSA THICKNESS

For Internal Use Only

WELFARE

MURAMIDASE DECREASES WET LITTER & FPD

84,9

97,9

14

2,121,07

75

80

85

90

95

100

NC BOND

FPD score (%) day 36

Score 2

Score 1

Score 0

a b

FPD score = 9.12 a 1.08bEsteve & Perez Calvo, 2019

2

CTR M007

CTR M007

For Internal Use Only

Increases Weight Gain (3%)

Improves digestibility of nutrients

Reduces thickness of GIT mucosa

Increases breast meat yield

Less wet litter and FPD

Improves FCR (-3%)

Increase absorption capacity

MURAMIDASE IMPROVES GASTROINTESTINAL FUNCTIONALITY

Muramidase

For Internal Use Only

There are a lot of birds out there

with their gut full of peptidoglycans

What are you going to do about it?

#FEED4GAIN

TAKE HOME MESSAGES

BRIGHT SCIENCE. BRIGHTER LIVING.

For Internal Use Only

#FEED4GAIN

TAKE HOME MESSAGES

Game Changing Innovation

Unlocking the Hidden

Potential

Novel and Unique

Muramidase

PGNs impact nutrient

absorption

Supporting gastrointestinal

functionality

Sustainable and environmentally friendly solution