Campylobacter and Salmonella in meat and eggs: the risk ... · Campylobacter and Salmonella in meat...

Campylobacter and Salmonella in meat and eggs: the risk from farm to forkand eggs: the risk from farm to fork

Marianne CHEMALY

Anses, UHQPAP: Hygiene & Quality of Poultry and Pork Products

Reported notification rates of zoonoses in confirmed human cases in the EU, 2011

(EFSA, 2013)

Trend in reported confirmed cases of human campylobateriosis in the EU, 2008–2011

(EFSA, 2013)

significant (p <0.001) increasein the last four years (2008–2011)

Trend in reported confirmed cases of human salmonellosis in the EU, 2008–2011

(EFSA, 2013)

Salmonella and Campylobacter

� Common reservoir: intestinal tract of a wide range of domestic and wild animals

� Contamination of foodstuffs by faecal organisms

� Transmission: introduction in food preparation

� Multiplication in food:• Inadequate storage temperatures• Inadequate cooking • Cross contamination

� Contamination: direct contact with infected animals or humans

� Campylobacter spp. most commonly associated with

human infection: C. jejuni followed by C. coli

� Chicken reservoir (80%)

� Broiler meat (20 to 30%): undercooked – cross

Salmonella and Campylobacter

� Broiler meat (20 to 30%): undercooked – cross

contamination

� Most frequent serovars: S. Enteritidis, S.Typhimurium

� Human S. Enteritidis cases: consumption of

contaminated eggs and poultry meat

Campylobacter and poultry meat

Epidemiological investigationsPrevalence, RF, molecular typing

Campylobacter : from farm to fork

Farms slaughterhouses ConsumersRetail

Host-pathogen interactionsVirulence, transfer, colonization

Control measuresFeed additives, chilling, …

Risk analysisQRA, Food safety criteria (PO, FSO, MC)


Campylobacter : prevalence and RF

72 % Caeca : 77 %Carcasses : 88 %Correlation

76 %Carcasses : 90 %Legs : 85 %Fillets : 53 %

Transfer rate: Up to 50 %Interaction time and weightFillets : 53 % weight

8 log10 CFU/g 2,4 log10 CFU/g < 2 log10 CFU/g

C. jejuni, C. coli

* O. Hue, S. le Bouquin, M.J. Laisney, V. Allain, F. Lalande, I. petetin, S. rouxel, S. Quesne, P.Y. Gloaguen, M. Picherot, J. Santolini,

S. Bougeard, G. Salvat, M. Chemaly. 2011. Campylobacter contamination of caeca and carcasses at the slaughterhouse and

correlation with Salmonella contamination. Food Microbiology. 28, 862-868.

* O. Hue, S. le Bouquin, M.J. Laisney, V. Allain, F. Lalande, I. petetin, S. rouxel, N. Homo, S. Quesne, P.Y. Gloaguen, M. Picherot, J.

Santolini, G. Salvat, S. Bougeard, M. Chemaly. 2010. Prevalence and risk factors for Campylobacter spp. contamination on broiler

chicken carcasses at the slaughterhouse. Food Microbiology. 27, 992-999.

* Huneau-Salaün A., Denis M., Balaine L., Salvat G. 2007. Risk factors for Campylobacter spp. Colonization in french free-range

broiler chicken flocks at the end of the indoor rearing period. Preventive Veterinary Medicine, 34-48.


Hot Season

PFs : water treatment, farm

practices

Season - Age

ThinningEvisceration: Dirty marks RoomT°°°°C

products with skinmore contaminatedthan productswithout skin(p<0.01) .

Campylobacter : prevalence and RF

practices RoomT°°°°C

PF : 1st batch

Correlation: carcass

contamination

significantly higher

(p<0.001) when

caeca are positive

(p<0.01) .

products packagedwith air greatercontamination(p=0.02) thanproducts packagedunder modifiedatmosphere.

High diversity: 0.978 PFGE

MLST


Campylobacter : molecular epidemiology

Farm: source infection to slaughterhousescross contamination

diversity

common STs : source of campylobacteriosis ST21 and ST 45

specific STs

* K. Rivoal, V. Rose, S. Quesne, F. Mégraud, M. Chemaly. 2013. A large scale survey describing the relationship between broilers

and human campylobacteriosis. CHRO, Aberdeen.

* K. Rivoal, S. Quesne, V. Rose, S. Rouxel, M. Chemaly. 2011. Prevalence of Campylobacter in retail broiler meat in France and

distribution of C. jejuni typed by MLST. CHRO, Vancouver.

Host-pathogen interactions: transfer


High prevalence

Transfer rate

?

Bacteriology ISO 10272Predictive microbiology

�Transfer may reach 50 %

�Contact time (p = 0.0095)

� Interaction weight and contact time (p = 0.0412)


�Transfer occurs even with very low load

� Transfer may occur even when contamination loadis below the detection limit

* Fravalo P., Laisney MJ, Gillard MO, Salvat G., Chemaly M. 2009. Campylobacter transfer from naturally contaminated chicken

thighs to cutting board is inversely related to the initial load. Journal of Food Protection. 72:1836-1840.

* Chemaly M., Laisney M.-J., Fravalo P. 2007. Modelling the transfer rate of Campylobacter from chicken thighs to kitchen cutting

surfaces. CHRO. V 54, suplement 1, p. 134.

� C. jejuni et C. coli are able to transfer (p > 0.05)

� Genetic diversity among isolates able to transfer

� Moderate to high adherence to inert surfaces

� All the strains are able to adhere and invade human

cells


cells

� C. jejuni more adherent and more invasive than C. coli

(p<0.05)

� Risk management

* Guyard M., Tresse O., Houard E., Jugiau F., Courtillon C., El Manaa K., Laisney M.J., Chemaly M. 2013. Characterization of

Campylobacter spp. transferred from naturally contaminated chicken legs to cooked chicken slices via a domestic cutting board.

International Journal of Food Microbiology. 164, 7-14.

Isolates


Host-pathogen interactions: virulence

Isolates

Virulence factorsPCR : LOS typingLOS A +B : orf7a/b : cstII

LOS B : orf5IIb : cgtA-IIb

LOS C : orf14c : glycosyltransférase

* M. Guyard, I. Kempf, E. Houard, S. Quesne, S. Rouxel, M. Chemaly. 2011. Correlation between sialylated Lipoologosaccharide

and antimicrobial resistance of Campylobacter jejuni isolates. CHRO, Vancouver.

50

60A

4% B21%

C

otherClasses

51%


Campylobacter: virulence

0

10

20

30

40

50

% d

es

is

ola

ts

Volailles

Humaines

A

Poultry

Humans

B C other classes

(n= 178)

(n= 150)

C24%

51%

A 9%

B 27%

C 29%

other Classes

35%

* M. Guyard, I. Kempf, E. Houard, S. Quesne, S. Rouxel, M. Chemaly. 2011. Correlation between sialylated Lipoologosaccharide

and antimicrobial resistance of Campylobacter jejuni isolates. CHRO, Vancouver.

Control measures

Biosecurity

Feed additives


Logistic slaughter

Chilling parameters

Regulation

Safety criteria

� product tests in vitro and in vivo

Feed additives

Vaccination

• Isolators• Animal husbandries• On-field

Chilling parameters Safety criteria

Education

Food authoritiesrecommendation

Feed additives In vitro In vivo

LAB -

Lactoserum based Bacteriostatic ?

Formic acid -

DL-methionine -

DL-HMTBA -

Plant extracts -

Humic acids -

Control measures

Humic acids -

Alguae Inhibition -

Organic acids Inhibition

Locust bean based -

Essential oils+organic acids -

Ion exchanged clay inhibition

Guyard M., Benzoni G., Quesne S., Bouvet D., Briant J., Guyonvarch A., and Chemaly M. 2013. Control of Campylobacter jejuni in

broilers using a feed additive. CHRO, Aberdeen.

Chemaly M., Lalande F., Quéguiner M., Fravalo P. 2007. The effect of a lactoserum-based product on the shedding of

Campylobacter by young turkeys. CHRO. V 54, suplement 1, p. 139.

Chemaly M., Lalande F., Quéguiner M., Fravalo P. 2007. The effect of an acid organic-based product on the shedding of

Campylobacter by young turkeys. CHRO. V 54, suplement 1, p. 139.

Data collectionAppropriate methods

Collaborations


Risk Analysis

CollaborationsBudget

Risk Analysis: safety criteria

Transport

Improving of fit-for-purpose sampling procedures for Campylobacter

Animal Receiving & Ante-mortem

Inspection

Stunning

Scalding

Plucking

Evisceration

Washing Chilling

Portioning, deboning

Processing

Packing, labelling

Transport

PO

FSO, MC

Risk Analysis: microbiological criteria and QMRA

� French Ministry of Agriculture and RA dpt in Anses

n = 5 c = 3 m M

Salmonella and eggs

How the eggs become infected?

oral / fecal cycle

Multiplicationfecal shedding

Digestive tract= non invasiveHorizontal transmission

MAJORITY OF SALMONELLA

SEROVARS

InvasionSeroconversion (Ab)

How the eggs become infected?

Organ infection(liver, ovaries,…)

Vertical Transmission (and horizontal)

SALMONELLA ENTERITIDIS

Consequences: layer production

� Hatchery

VERTICALHORIZONTAL

� Impact on production stages

Consequences: public health

• Survival during storage• absence of multiplication• Outbreak if multiplication (storage T°°°°C)

� Eggs and consumer

Horizontal

• Outbreak if multiplication (storage T°°°°C)

• Access to yolk• yolk = broth culture• Multiplication and outbreak(even well handled)

Vertical

Regulation laying hens

�Since 1992: S. Enteritidis and S.Typhimurium (not mandatory)

� EU regulation 2003 : S. Enteritidis - S.� EU regulation 2003 : S. Enteritidis - S.Typhimurium – S. Hadar – S. Virchowand S. Infantis (mandatory)

Regulation eggs and egg products

� MC (EC N°1441/2007)

� Enterobacteriacea

� Italy

PO: after packaging (finished product)

FSO: retail (0-28days)

� Croatia

Prevalence LH flocks below 1%� n = 5 c = 2

�m = 10 ufc/g or ml M = 100 ufc/g or ml

� End of the manufacturing process

� Checks on the efficiency of the heat

treatment and prevention of

recontamination

Prevalence LH flocks below 1%

absence of Salmonella in 25 g (whole eggs)

(1/1000 tons of annual egg production

retail (0-21 days)

�France

Control of laying hen flocks for SE

Egg labeling (tracking)

� S. Enteritidis

� Common reservoir: gut

� Transmission: direct contact, cross contamination, inadequate storage, etc.

At the farm

� EU baseline study in 2004: • 0 to 79.5 % S. spp.•18.4 % S. Enteritidis

� RF: size, housing, season, age

Huneau-Salaün A., Chemaly M., Le Bouquin S., Lalande F., Petetin I., Rouxel S., Michel V., Fravalo P., Rose N. 2009. Risk factors

for Salmonella enterica subsp. enterica contamination in 519 French laying hen flocks at the end of the laying period. Preventive

Veterinary Medicine. 89:51-58.

At the farm

� Vertical or horizontal infection

� Vertical: scarce (1/14040)

� Horizontal: rearing, packing

Eggshells

� Rearing: 39 % of + flocks lead to positive eggs

� Packing: cross contamination, C&D

� RF: size, environmental conditions, laying rate, age

Chemaly M., Huneau-Salaun A., Labbé A., Houdayer C., Petetin I., Fravalo P. 2009. Isolation of Salmonella enterica in laying hen

flocks and assessment of eggshell contamination in France. Journal of Food Protection.72:2071-2077.

PFGE-Xba1+PFGE-Bln1

Xba1-Bln1

10

0

80

60

40

PFGE-Xba1 PFGE-Bln1

S. Enteritidis egg (335)

S. Enteritidis farm (335)









S. Enteritidis

� Genetic characterization of isolates (flocks – eggs)

Eggshells


S. Montevideo egg (240)

S. Montevideo farm (240)

S. Infantis egg (475)

S. Infantis farm (475)

S. Virchow egg (486)

S. Virchow farm (486)



S. Typhimurium egg (359)


S. Typhimurium farm (359)


• 22 isolates: 11 flocks / 11 eggs• 10 S.E. – 4 S.T. – 4 S.V. – 2 S.I. – 2 S.M.

S. Montevideo

S. Virchow

S. Typhimurium

S. Infantis

PFGE-Xba1+PFGE-Bln1

Xba1-Bln1

10

0

80

60

40

PFGE-Xba1 PFGE-Bln1











100 % similar

S.E.: 85 %

� Genetic characterization of isolates (flocks – eggs)

Eggshells


S. Montevideo egg (240)

S. Montevideo farm (240)

S. Infantis egg (475)

S. Infantis farm (475)









S.V.: similar/flock

S.T.: 1 isolate 90 %

• 22 isolates: 11 flocks / 11 eggs• 10 S.E. – 4 S.T. – 4 S.V. – 2 S.I. – 2 S.M.

� Identification of factors associated to the presence of Salmonella oneggshells

Results

� 39 % positive flocks: positive eggs

� Frequency of contamination of eggs: 0.6 to 8.6 %

� Flocks and eggs: same serovars – similar isolatesand/or very close

� Associated factors:

Eggshells

� Associated factors: • Environnemental contamination • Flock size• Rearing practices: other productions

� Microbiological quality of eggs

� Salmonella may penetrate

� Potential sources for egg dishes contamination

Egg products

� S. Enteritidis: mostly involved in outbreaks

� Growth control: • pasteurization (time x T°°°°C)• salt, suggar, lysozyme, pH

� QRA: 1763, 407 and 0 cases (0, 10% salt, 10% sugar)

S. E. : sweet matrix

Factor Effect p

T°C + (↑) 0.000 R = 0.97

Egg products

Sugar - (↓) 0.0013 R2 = 0.93

T°CxSugar - (↓) 0.0023 LoF = 0.06

SucrexpH - (↓) 0.0359

S. E. : salty matrix

Factor Effect p

T°C + (↑) 0.0025 R = 0.94

Egg products

Salt - (↓) 0.0016 R2 = 0.83

T°CxSel - (↓) 0.0031 LoF = 0.051

QRA and existing models

� Many QRA since 1997 (Buchanan)

� S.E. in shell and liquid eggs

� FSIS: model at national level in 1998

• Infection related to eggs internally contaminated: incidence of

illness

� other models: contamination of eggs, risk of exposure, efficiency of

intervention strategies, etc.

� WHO/FAO (2002) and EFSA (2010)

Schematic diagram showing the stages of the risk assessment for Salmonella in eggs


� Major RF :

• Within flock prevalence (prevalence � risk �)

• importance of intervention strategies to control SE

(biosecurity measures, egg pasteurization,…)

• internal infection: low amount, low risk but X during

transport, storage and handling

� EFSA model (2010):

• Two stage bayesian model

• N° of contaminated eggs (ext. and int.) by SE

• 1st: average flock prevalence over a laying period

• 2nd: estimation of proportion of + eggs within an infected flock

• estimation at a national level the average contamination per million


� EFSA opinion (2010): « quantitative estimation of the public health impact of setting

a new target for the reduction of Salmonella in laying hens”

• linear relationship: flock prevalence and infected eggs

• lack of data: N° infected eggs within an infected flock

� Current WG (2013-2014)

�Safety of poultry products constant progress for Salmonella

� Improve biosafety measures: recontamination by otheranimal productions

�Solutions to find forCampylobacter reduction in poultry

Conclusion

�Solutions to find forCampylobacter reduction in poultry

� Improve process practices (evisceration…) at the slaughterhouses

�Many efforts to improve consumer education / information

� Implementation of regulation

Conclusion

Campylobacter and Salmonella in meat and eggs: the risk ... · Campylobacter and Salmonella in meat...

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