Florence Baron – Sophie Jan

Agrocampus Rennes
France

Egg
Egg and
and egg
egg product
product microbiology
microbiology ::
actual
actual consideration
consideration onon breeding
breeding
and
and technological
technological practices
practices

1
 I-Microbiology
I-Microbiology of
of eggs
eggs

Egg
Egg contamination
contamination
Exogenic contamination

Egg content is
Penetration
generally sterile…
but not the shell

endogenic contamination
(ovary)

Multiplication in the egg

compartments

2
 I-Microbiology
I-Microbiology of
of eggs
eggs

Endogenic contamination of eggs

Salmonella enterica serovar Enteritidis , Heidelberg, pullorum, galinarum

(Van Immerseel et al., 2005 ; Gast and Board, 1992)

Campylobacter (Sahin et al., 2004)

Ovary Contamination of egg yolk or
vitellin membrane
Hen : Normal host
of Salmonella and
Campylobacter Magnum Contamination of egg white

Strong affinity for usually

the genital tract Isthmus
Contamination of egg shell
Uterus membranes or egg shells
Vagina Gast and Beard 1990, Humphrey 1994,
Hoop and Pospischil 1993,
Miyamoto et al. 1997,
Okamura et al. 2001 3
 I-Microbiology
I-Microbiology of
of eggs
eggs

Exogenic contamination of eggs

Microorganisms from :
- the digestive tract of the hen
- the breeding environment

The level and type of contamination vary according to the study,

the breeding and the geographical area :
- From 103 to 106 bacteria/egg

- Prevalence of G+ cocci :
Micrococcus, Aerococcus, Streptococcus, Staphylococcus, etc.
- Presence of enterobacteria (G-) :
Enterobacter, Escherichia, Salmonella, Citrobacter, Klebsiella, Serratia, etc.

- Presence of Bacillus Board 1969, Moats 1980, Mayes and Takeballi 1983,
Wood and Waites 1988, Musgrove et al., 2005
4
 I-Microbiology
I-Microbiology of
of eggs
eggs

Exogenic contamination of eggs

The level of contamination is influenced by :
- the level of air contamination
- the season (summer > winter)
- the system of breeding and cage design
It is difficult to compare the studies :
- No differences between conventional cages and furnished cages but
differences between cages and aviary housing systems (De Rheu et al., 2005)
- Differences between conventional cages and furnished cages (Mallet et al., 2005)
Depends on the way in which the eggs are collected

¨ Low contamination of eggshells for all the systems

Importance of egg localization (nest box, dust bath, litter) on the number of
dirty and/or cracked eggs. The use of perches increases the number of
cracked eggs
Work on the design of the cages/the behaviour of hens
5
 I-Microbiology
I-Microbiology of
of eggs
eggs

Penetration
Natural barriers against penetration:

Cuticule :
- prevents the microorganism penetration through the
shell pores
- fragile

Eggshell :
perforated thick barrier :
physical barrier but not very effective

Eggshell membranes :
- anti- bacterial « filter » with bacteriostatic properties
- effective barrier

6
 I-Microbiology
I-Microbiology of
of eggs
eggs

Penetration
Factors influencing the penetration :

- Maturity of the cuticule just after the laying

- Temperature changes
- Condensation (increases the penetration without impacting
on the contamination of the content) De Rheu et al. 2005
Importance of storage
Difficult to make the distinction between shell or shell membrane 40°C
contamination during egg formation and contamination of egg surface
at the time of the passage in the cloaca follow-up of a penetration

20°C
In the case of penetration : selective effect (De Buck et al., 2004)

No influence of the quality of the shell (thickness, rigidity,

number of pores)

No influence of the breeding system (Messens et al., 2005)

7
 I-Microbiology
I-Microbiology of
of eggs
eggs

Glucids Minerals
Migration-multiplication in egg white protids (0.6%) (0.58%)
Lipids
(10.8%) (0.024%)
Non-favourable environment for bacterial growth
¨ Low growth for some microorganisms
¨ Bacteriostatic or bactericide effect for others Water (88%)

Strong selective effect .

- alkaline pH : bacteriostatic, flagella ? But not just after laying
- viscous : retards the migration Time-dependent decrease in viscosity
- iron deprived
- many inhibiting proteins :
Garibaldi, 1970
- Lysozyme Nath and Baker, 1973
Foegeding and Stanley, 1987
- Ovotransferrin
Bradshaw et al., 1990
- Protease inhibitors Lock and Board, 1992
Baron et al., 1997
- Proteins chelating essential elements Gast and Holt, 2000
- + molecules and/or unknown systems Messens, 2004
8
 I-Microbiology
I-Microbiology of
of eggs
eggs

From egg white to egg yolk

Egg white : Non-favourable environment for bacterial growth

+ Chalazes
maintain the yolk in central position :
barrier of structure Decreases as a function of preservation time

Importance of storage
9
 I-Microbiology
I-Microbiology of
of eggs
eggs

Multiplication in egg yolk

Contains growth elements
Egg yolk composition :
for embryo development
Glucids
Lipids (1 - 2%)
(32 - 36%)
Ideal for microorganism
development
Water (50%)
Ex : inoculation of egg yolk with 20 cells
Protids of SE /mL : 109 cells /mL within less
(16%) than 24 h at 23°C (Saeed and Koons, 1993)

Iron rich, vitamin rich, etc.

Importance of the
storage temperature

10
 I-Microbiology
I-Microbiology of
of eggs
eggs

Whole egg : the cuticule, the shell, the shell

membranes, the egg white, the chalazes prevent
the penetration, the multiplication and the
progression of the microorganisms towards the egg
yolk where they could reach important
concentrations !

But :
- Microorganisms can also set up systems enabling them to deviate
certain defenses of the egg.

- The effectiveness of egg defenses can decrease in case of bad

handling, bad preservation, etc.

11
 I-Microbiology
I-Microbiology of
of eggs
eggs

How to reduce egg contamination ?

Caution :
¨ At the breeding level : often empirical employment,few scientific data
Consequences ?
- Reduction of the poultry digestive tract colonization by Salmonella :
control, destruction, vaccination (Nassar et al., 1994 ; Holt et al., 2003), food
additives (Van Immerseel et al., 2004 ), prebiotics (Bailey et al., 1991), probiotics
(Mulder et al., 1997 ; Pascual et al., 1999)

- Reduction of the contamination of the breeding environment : hygiene

procedures (cleaning and disinfection), “anti-Salmonella” positive
biofilms (Lactobacillus, Pediococcus, Bacillus)

¨ At the storage level :

- Temperature, duration, hygiene

¨ At the conditioning level :

- Control (check - candling), egg decontamination 12
 I-Microbiology
I-Microbiology of
of eggs
eggs

Egg decontamination procedures

¨ Washing of table eggs (category A)
€ prohibited in Europe
€ authorized in the USA, in Canada, Japan, Australia, Russia, Mexico,
Saudi Arabia and Sweden
Positive arguments Negative arguments
Reduction in the surface Musgrove et al., Damage of natural defenses Kim and Slavik, 1996
microbial load 2004 and 2005 (especially the cuticule) by Wang and Slavik, 1998
(from 1 to 6 log10) physical and chemical agents Favier et al., 2000

Lucore et al., 1997 Haines and Morand,

Hutchison et al., 1940
2003 and 2004 Lorenz and Starr, 1952
Reduction in the internal Jones et al., 2004
Increase in the internal Brant and Starr, 1962
microbial load microbial load

Caution :
No epidemiological data on the consequences of egg washing
on public health !
13
 I-Microbiology
I-Microbiology of
of eggs
eggs

Opinion of the Scientific panel on Biological Hazards on the request from the
Commission related to the Microbiological Risks on Washing of Table Eggs. The EFSA
Journal (2005), 269, 1-39
Use of chemical agents not damaging Dirty
Immediately after drying the cuticule and leaving no residues eggs

Scaling Check- Oiling Drying Washing Accumulation

candling

Control of water
temperature and quality

Cleaning of the whole circuit

Sorting of eggs
(+ water tanks) every 4 h Loading
before washing

Storage :
Packer head lanes ¨ < 7 days after laying
¨ below 8°C
¨ dry area
Adaptated from Musgrove et al., 2005 14
 I-Microbiology
I-Microbiology of
of eggs
eggs

Other egg decontamination procedures

¨ UV (Favier et al., 2001 ; De Rheu et al., 2005)

¨ Whole egg pasteurization (Denys et al., 2005)

¨ Ozone (Whistler and Sheldon, 1989 ; Rodriguez-Romo and Yousef, 2005 ; Davies and Breslin, 2003)

¨ Oxidized ionized water (Bialka et al., 2004)

¨ Ultrasounds (Cabeza et al., 2004 et 2005)

¨ New detergents (Protais et al., 1997)

Caution :
often on an experimental scale, discussed opinions
Consequences ?

15
 I-Microbiology
I-Microbiology of
of eggs
eggs

Conclusion
Egg contamination depends on :

Localization and Breeding practices collecting, washing and

level of Anti-Salmonella procedures storage conditions
contamination
Any change in the selection, the breeding practices, the process of
decontamination, the preservation, can potentially modify the
contamination, the penetration and/or the multiplication of the micro-
organisms 16
 II-Microbiology
II-Microbiology of
of egg
egg products
products

Egg
Egg contamination
contamination Egg
Egg product
product contamination
contamination

Labour Environment

Egg products
Methods Equipment

17
 II-Microbiology
II-Microbiology of
of ovoproducts
ovoproducts

Once broken,the egg loses its natural defences :

- Whole egg and egg yolk : ideal environment for microorganism

development
- Egg white : non-favourable environment : low growth,
maintenance or decrease in microorganism population

How to control egg microbiology ?

- Egg quality
- Respect of good practices
- Transformation-stabilisation

18
 Reception and storage of eggs Diagram of egg product processing
Breaking
Eggshells

Whole egg Egg yolk Egg white

Filtration
Egg white
Cooling
Concentration
Dry egg products Sugar removal
Intermediate storage
(sugaring, salting, etc.) (powders)
Drying
Liquid Pasteurization
egg products Whole egg Egg yolk Conditioning

Cooling
hot room
Drying
(pasteurization)
Conditioning
Conditioning Cooling

Storage Congelation
Storage Storage

Delivery
19
 Reception and storage

Breaking
Eggshells
Reception :
Controls,
Whole egg suppliers
Egg yolk Egg white

% of dirty or cracked eggs

Filtration
Egg white
Cooling
Storage : Concentration
Temperature, duration,
Intermediate storage Dry ovoproducts Sugar removal
(sugaring, salting, etc.) (powders)
Recycling cells (plastic / paperboards) Drying
Liquid Pasteurization
egg products Whole egg Egg yolk Conditioning

Cleaning and disinfection

Cooling of the shells :
hot room
Authorized in egg breaking factory (European directive, Drying
1989)
(pasteurization)

In practice Conditioning
in France : some producers still begin… Improvement
Conditioning needed
Cooling
of the control of the process
Storage Congelation
Storage Storage

Delivery
20
 Reception and storage

Breaking Egg shells

Whole egg Egg yolk Egg white

Filtration
Egg white
Cooling
Concentration
Dry egg products Sugar removal
Intermediate storage
- Effective separation
(sugaring, salting, etc.) (powders)
Drying
- Fast elimination of the shells
- Shell debris
Liquid Pasteurization
ovoproducts Whole egg Egg yolk Conditioning

Cooling
hot room
Drying
(pasteurization)
Rotary breaker
Conditioning On-line breaker
Conditioning • Importance of the adjustment
Cooling
of the machines and human
Storage Conglelation monitoring
Storage Storage
• Respect of good practices
Delivery
21
 Reception and storage

Breaking
Eggshells

Whole egg Egg yolk Egg white

Filtration Elimination of shell debris

Egg white
Cooling
Concentration
Dry egg products Sugar removal
Intermediate storage
(sugaring, salting, etc.) (powders)
Drying
Liquid Pasteurization
Caution : difficulties
Whole egg to Egg
clean
yolk the filter
egg products Conditioning

Cooling
hot room
Drying
(pasteurization)
¨ Remaining shell debris
Conditioning
Conditioning
¨ Biofilm development Cooling

Storage ¨ egg
Congelation product contamination
Storage Storage

Delivery
22
 Stabilisation (1 Separation 2 Inhibition 3 Destruction)
Reception and storage
Jeantet and Baron 2003

1 Separation ofBreaking
microorganisms from the product : microfiltration and centrifugation
Eggshells

2 Decrease in the growth kinetics of microorganisms : cooling, acidification,

Whole egg Egg yolk Egg white
inhibitor addition (sorbic and benzoïc acids), decrease in aw (concentration,
drying, congelation)
Filtration
Egg white
Cooling
L Psychrotolerant Concentration
Concentration
species during storage ! Dry egg products Sugar removal
Sugar removal
Intermediate storage
(sugaring, salting, etc.) (powders)
Drying
Liquid Pasteurization
Pasteurization
eggproducts
ovoproducts Whole egg Egg yolk Conditioning

Cooling
stoving
hotinroom
hot room
Drying
(pasteurization)
(pasteurization)
Conditioning
Conditioning Cooling

Storage Congelation
Congelation
Storage Storage
3 Destruction of microorganisms : thermal treatments (pasteurization), ionization
(decree of October 1990), addition of Delivery
bactericidal agents (Nisin), PFGE (Martin-
Belloso et al. 1997, Jeantet et al. 1999) 23
 Stabilisation (1 Separation
Reception and storage 2 Inhibition 3 Destruction) Jeantet and Baron 2003

Breaking
Eggshells

Wholeegg
egg Eggyolk
yolk Eggwhite
white
Non effective for spores
Whole Egg Egg 55-57°C (2-6 min) or heat-resisting species
65 à 70°C (2-6 Filtration
min)
Egg white
Cooling
Concentration
Dry ovoproducts Sugar removal
Intermediate storage
(sugaring, salting, etc.) (powders)
Drying
Liquid Pasteurization
Pasteurization
egg Conditioning
Whole egg Egg yolk
products
Cooling
stoving in hot room
Drying
(pasteurization)
Conditioning
Conditioning Cooling

Storage Congelation
Storage Storage
3 Destruction of microorganisms : thermal treatments (pasteurization), ionization
(decree of October 1990), addition of Delivery
bactericidal agents (Nisin), PFGE (Martin-
Belloso et al. 1997, Jeantet et al. 1999)
24
 Stabilisation (1 Separation
Reception and storage 2 Inhibition 3 Destruction) Jeantet and Baron 2003

Breaking
Eggshells
LThe effectiveness of egg defenses can
decrease with the transformation process
Whole egg Egg yolk Egg white

Ex : Pasteurization of egg white powder

Filtration (Baron et al., 2003) :
Egg white
Loss of the Cooling
bacteriostatic
capacity of reconstituted egg Dry egg products Concentration
white with respect
Intermediate to Salmonella
storage Dry(powders)
eggproducts Sugar removal
(powders)
¾ due to ovotransferrin denaturation
(sugaring, salting, etc.)
Drying
Pasteurization at at
Pasteurization
¾ Liquid 67°C 67°C : maintenance of
bacteriostatic potential, improvement ofEgg yolk
foaming
Pasteurization
ovoproducts Conditioning
Whole egg
properties while no effect on gelling properties
Cooling
Pasteurizationat at
¾ Pasteurization 75°C 75°C : very effective for hot room
improvement of gelling properties, no additional profit
Drying
(pasteurization)
as compared toConditioning
67°C for the foaming properties and
strong decrease in the bacteriostatic properties
Conditioning Cooling

Storage Congelation
Storage Storage
3 Destruction of microorganisms : thermal treatments (pasteurization), ionization
(decree of October 1990), addition of Delivery
bactericidal agents (Nisin), PFGE (Martin-
Belloso et al. 1997, Jeantet et al. 1999)
25
 Reception and storage 103-4 Level of contamination during the process
Baron et al., unpublished
Breaking
Egg shells 106-8
Whole egg Egg yolk Egg white 102-3

Filtration
104 Egg white
Cooling

106
Concentration
Dry egg products Sugar removal
Intermediate storage
(sugaring, salting, etc.) 105 (powders)
Drying
Liquid Pasteurization
egg products Whole egg Egg yolk Conditioning
102
Cooling
Drying
hot room
(pasteurization)
<10
Conditioning
Conditioning Cooling

Storage Congelation
Storage Storage
102
Delivery
26
 II-Microbiology
II-Microbiology of
of ovoproducts
ovoproducts

Which microorganisms contaminate the egg products ?

- Selective effect of the egg product
- Selective effect of the preservation-transformation processes
Exhaustive study on whole egg before and after pasteurization
Pôle Agronomique Ouest – Bretagne and Pays de Loire (France)
(3 egg breaking manufacturers + 7 laboratories : AFSSA-Ploufragan, ADRIA
Quimper, Agrocampus Rennes, ASEPT-Laval, ENV-Nantes, ESMISAB-Brest,
Nutrinov-Rennes)

Before pasteurization After pasteurization

Salmonella ++++++ (+)
Listeria + -
Campylobacter ++ -
Treatment effective on pathogenic species
Seasonal effect : - Autumn : lower contamination levels
- Summer : higher contamination levels
(especially psychrotroph species)
- Winter : Streptococcus, Enterococcus
27
 Conclusion
Conclusion

Collecting, Flora of
Breeding practices washing and table eggs
Anti-Salmonella procedures storage

Whatever the step, any +

modification can modify egg
and egg product flora

It is essential to Flora of egg

consider the effects Transformation, products
of any modification stabilisation
28