Effect of drinking water additives on growth performance and

intestinal microbiota composition in weaned piglets

Juan Antonio Mesonero Escuredo, DVM

Resident ECPHM

Global Programme Manager Swine Gut Health

Content

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Introduction to the chemical basis for intestinal health

Pharmacology and pH

Water quality basis for Intestinal health

Implementing additive strategies in Australia and EU

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1.Chemical basis of organic chemistry for intestinal health

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pH and pKa index

pH

pKa, dissociation index

A-COOH + H2O A-COO- + H3O+

Chemical basis, the pillars

1. Organic acids (OAs) vs. Inorganic acids2. Molecular weight, molecular size, the concentration, number of active particles3. pKa value & pH of the environment and temperature4. Relative lipophilicity as indicated by its Log Pow

a) The bacterial surface area, permeability and thickness of membrane and bacterial wall.

5. Buffered vs. no buffered, application of salts of buffered OAs (e.g., ammonium formate) can deliver an anion for OAs production at a low pH, such as that present in the stomach.

6. SCFA vs MCFA7. Spectrum of action

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Antibacterial mode of action of organic acids

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- In an attempt to expel C8 and C10 and to restore the pH the cell consumes enormous amounts of energy that lead to cell death. - In the case of C10 the excessive production of reactive oxygen species (ROS) leads to additional cell damage

C8 80 %20 %

Outside the cell Inside the cellcmcw

acid neutral

C10 30 %70 %

ethyl-decanoate

Tpo1pdetox

H+

protonpump

H+

detox

ATP

ADP

Pdr12pC8

H+ H

+Tpo1pperoxisome

activated fatty acid β-oxidation

ROS (H2O2)

cell death

Toxicity mechanism of C8 and C10 in S. cerevisiae

Mode of action of organic acids

In addition, for solute ionizing substances;

1. The membrane charge

2. Transmembrane ionic gradient and possibility that the solute may be a substrate of a carrier protein must be considered.

3. Many OA are substrates of the monocarboxylate transporters (Sepponen K, et al. 2007).

4. Recent studies showed that the mode of action of organic acids in bacteria, yeast and molds is related to dissipation of the proton gradient (“classical” uncoupling effect), acidification of the cytosol, membrane disruption, osmotic stress by anion accumulation and metabolic disturbance by inactivating specific enzymes.

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Basis of pharmacology related to pH and pKa

Pharmacokinetics - pKa and drug absorption

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The environmental pH and the individual pKa value determine the pharmacokinetics of the compounds

pKa values of common drugs

Pharmacokinetics and Pharmacodynamics

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Influence of pH in molecules absorption (stomach example, weak acid)

2.Water quality basis for intestinal health

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Water quality basis for intestinal health1. Water pH and pKa index

2. Water hardness

3. Water Oxidative- Reduction Potential (ORP)

4. Water microbiology

5. Water Total Dissolved Solids (TDS)

6. Water chemistry

7. Water antibiotics and vaccines

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Water microbiology

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Reference water <1000 <100 <100

3.Implementing drinking additive strategies

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1. Australia AB reduction case study

2. Swine Research Center, SRC (The Netherlands)

3. Salmonella SRC & Field OD case studies

Strategies vs. field challenges

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Specie Segment Value Standard Strategy

Swine Sows Water consumption Continue use

Infection pressure sow-piglet 2-4 weeks before farrowing

Urinary issues Continue use

Lactation, body condition Continue use during lactation

Piglets Mortality, diarrhea Continue use the first 4 week after weaning

FCR, ADG, Uniformity Continue use

Antibiotic replacement Continue use in the target periods

Finishers Salmonella control Continue use

FCR Continue use

Mortality Continue use

Antibiotic replacement Continue use in the target periods

1. Continue use means 24 hours per day, 7 days per week2. All strategies are reviewable case per case

Australia

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• Journal of Porcine Health Management, 2016

Material and methods

420 weaned piglets (21 days) 3 treatments (Full AB (Amoxicillin, Neomycin), Full AB + OAs, Reduced AB (Amoxicillin)

& OAs) 10 pens per treatment during 4 weeks post-weaningM. hyopneumoniae and APP (Serovar 15) positive PCV2 positive

H. parasuis positive E.coli O139:K88 positive. In the past O:141 and O:149

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Water titration curve

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OAs improve LW, ADG and FCR in weaned piglets

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4

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Entry Week 1 Week 2 Week 3 Week 4

aa

b

Live

we

igh

t (k

g)

OAs improves live weight

Control

OAs + full medication

OAs + reduced medication

13,5 Kg.

12,5 Kg.

12,8 Kg.

0

0.5

1

1.5

2

ADG ADFI FCR

a

a

a

a

b

b

OAs improves average daily gain and FCR

Control

OAs + full medication

OAs + reduced medication

-17.0%

+16.2%

Economical impacts

Improved final weight by 1 Kg in 4 weeks, even with reduced medication.

Better feed conversion ratio by 0,3 points, even with reduced medication.

Higher water usage in the two groups using additives.

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The Netherlands(EU)

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• 9th European Symposium of Porcine Health Management (ESPHM), 2017

Material and methods

o 120 weaned piglets, 28 old days 2 treatments, 3 animals per pen, 20 replicates per treatment

Animals were blocked by body weight and sex, blind study. 6 week study period Feed challenge with high CP (20.6%) and high Ca (5%, 8%, 8% phase1, phase2, phase3)

o Severe diarrhea and switch to commercial feed on day 29 of study (ethical decision) Weekly BW & FI, daily water intake & diarrhea scores At the end of the study 10 animals per treatment

o Stomach and gut pHo Microbiota analysis

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Group Treatment Test article Test article inclusion

1DWA Blend of OAs; Formic, Acetic and Ammonium Formate (Selko-pH)

2.0 l/m3

Water Titrated2

Control Water -

DWA Positive effect on BW, ADG, ADF and ADW

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Treatment p-value

DWA Control treatment

n 20 20

BW (Kg) d 42 26.3a 24.4b <0.001

ADG (g/d) d 0-42 447a 402b 0.006

ADF (g/d) d 0-42 645a 568b 0.026

FE (ADG/ADF) d 0-42 0.715 0.707 0.695

ADW (ml/d) d 0-42 1937a 1630b 0.003

o DWA had significant effect on: Increased body weight and average daily gain Increased feed and water intake

DWA reduce water pH, no significant on stomach and gut

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Group Product Time dosed (days in study) Dose (ml/l) Water pH

1 DWA 0-42 2 3.5-3.7

2 Control 0-42 0 7.2-8.0

Results on gut and stomach pH

Treatment

DWA Control p-value

n 10 10

Stomach 2.7 3.4 0.372

Proximal 5.6 6.0 0.604

Distal 6.9 6.7 0.156

Results on gut and stomach pH

Significant reduction in the amount of Streptococcus

DWA Control

o Phylotyping (genus level)

Discussion & Conclusion

o Driking water additives based on Formic, Acetic and Ammonium Formate had significant effect:

Increasing body weight and average daily gain

Increasing feed and water intake

Shows to be a tool in AB reduction

Significant reduction in the amount of Streptococcus genus

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Salmonella

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• Journal of Animal Science, 2017

• Field experiences in The NL

R&D Fecal shedding of Salmonella after infection

Peak shedding 1.4 log CFU/gram lower with Selko-pH in drinking water (p=0.015)

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0 5 10 15 20 25

CFU

Sal

mo

ne

lla /

gra

m f

ece

s

Days post infection

non-infected

infected

Selko-pH

If we reduce Salmonella in our model with 1 log, we see a strong reduction in presence of Salmonella at the farms. Average OD-values from 40 to 10.

2 log reduction of meat contamination gives 1 log reduction of human salmonella cases

Field results of Salmonella reduction over 2 years

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Q1 2014 Q2 2014 Q3 2014 Q4 2014 Q1 2015 Q2 2015 Q3 2015 Q4 2015

Piglets

% positive 35 17 5 5 0

Average OD 48 23 11 9 0

Grower finisher

% positive 70 63 40 5 0 0 0 0

Average OD 99 87 57 27 19 29 27 18

Salmonella category 3 3 3 2 1 1 1 1

Diagnostic – water analyses* Q1 2014

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Acids used pH level Yeast (cfu/g) Moulds (cfu/g) Enterobacteria (cfu/g)

Piglets room 32 yes 4,44 > 100.000 3.000 < 100

Piglets room 34 yes 4,45 > 100.000 4.000 < 100

Piglets room 39 yes 4,41 15.000 1.000 < 100

Fatteners room 62 yes 3,70 1.000 1.000 < 100

Fatteners room 65 yes 3,66 < 100 < 100 < 100

Fatteners room 69 yes 3,64 < 100 < 100 < 100

Reference water <1000 <100 <100

*Orion quick dip slides

*Metrohm titrator

Diagnostic – feed analyses* Q1 2014

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Acids used pH level Yeast (cfu/g) Moulds (cfu/g) Enterobacteria (cfu/g) aw

Grinded Corn 6 hours old No 6,66 10.000 2.200 6.000 -

Grinded Corn 1 day old No 6,53 44.000 70.000 7.500 -

Piglet feed room 35 No 6,56 220.000 < 100 < 100 -

Fattener feed room 60 No 6,06 20.000 5.000 32.000 -

Start feed fattener No 5,96 50.000 1.500 4.000 -

CCM No 4,25 37.500 < 100 < 100 -

Fresh grinded wheat No 6,37 1.450.000 60.000 700.000 0,81

Wheat No 7,35 620.000 1.000 800.000 0,81

Reference < 100.000 < 10.000 <10.000 < 0,68

Feed 10.000 cfu/g Enterobacteria, 1 cfu/gr risk of Salmonella (Jones & Richardson, 2004)

*Feed and raw material analyses based on ISO methods

Solution

1. Feed management:

Feed preservation: 5 kg/mT (Fysal; formic, acetic, propionic, ammonium formate)

Continue follow-up of the potential source of Salmonella infection

2. Farm management:

Reminding and remarking strict hygiene, different areas, etc

Discussing with the employee the challenge & continue education

3. Health Management:

Blood samples following the Salmonella OD levels, where are they high now?

Water drinking additive strategy and blend changed

Drinking additives (Selko-pH; formic, acetic and ammonium formate) protocol for each production phase (limitation of water treatments in sows due to the water lines)

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Results of Salmonella reduction over 2 years

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Q1 2014 Q2 2014 Q3 2014 Q4 2014 Q1 2015 Q2 2015 Q3 2015 Q4 2015

Salmonella category

Other advantages• ADG improvement: 30 gram• Reduction FCR: 0,1• Lower feed costs• ROI: 3 (calculated based on fattening period)

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60

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Q1 2014 Q2 2014 Q3 2014 Q4 2014 Q1 2015 Q2 2015 Q3 2015 Q4 2015

Blo

od

OD

val

ue

OD values for piglets and fatteners

Average OD piglets Average OD fatteners

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Acknowledgements

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This presentation wouldn’t be possible without AUSPAC and TN R&D team contribution, thanks to all coauthors.

Christine ClarkAUSPAC, Australia

A.K. Oudshoorn, M. Davids, Paul BruinenbergP. Roubos-van den Hill, PhDTrouw Nutrition R&D, Boxmeer, Netherlands.

Dr. John Carr,Specialist Pig Practice, www.carrsconsulting.com, Melbourne, Australia.

Dr. Dominiek Maes,University of Ghent, Department of Reproduction- Obstetrics and Herd Health- Unit Porcine Health Management, Merelbeke, Belgium

Dank U!

Takk!

Merci!

ありがとう！

谢谢！