campylobacter and salmonella in meat and eggs: the risk ... · campylobacter and salmonella in meat...
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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)
Farms slaughterhouses ConsumersRetail
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.
Farms slaughterhouses ConsumersRetail
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
Farms slaughterhouses ConsumersRetail
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
Farms slaughterhouses ConsumersRetail
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)
Host-pathogen interactions: transfer
�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
Host-pathogen interactions: transfer
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
Farms slaughterhouses ConsumersRetail
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%
Farms slaughterhouses ConsumersRetail
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
Farms slaughterhouses ConsumersRetail
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
Farms slaughterhouses ConsumersRetail
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
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 egg (44)
S. Enteritidis farm (44)
S. Enteritidis egg (54)
S. Enteritidis farm (54)
S. Enteritidis egg (191)
S. Enteritidis farm (191)
S. Enteritidis farm (31)
S. Enteritidis egg (31)
S. Enteritidis
� Genetic characterization of isolates (flocks – eggs)
Eggshells
S. Enteritidis egg (31)
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. Virchow egg (357)
S. Virchow farm (357)
S. Typhimurium egg (359)
S. Typhimurium egg (389)
S. Typhimurium farm (359)
S. Typhimurium farm (389)
• 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
S. Enteritidis egg (335)
S. Enteritidis farm (335)
S. Enteritidis egg (44)
S. Enteritidis farm (44)
S. Enteritidis egg (54)
S. Enteritidis farm (54)
S. Enteritidis egg (191)
S. Enteritidis farm (191)
S. Enteritidis farm (31)
S. Enteritidis egg (31)
100 % similar
S.E.: 85 %
� Genetic characterization of isolates (flocks – eggs)
Eggshells
S. Enteritidis egg (31)
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. Virchow egg (357)
S. Virchow farm (357)
S. Typhimurium egg (359)
S. Typhimurium egg (389)
S. Typhimurium farm (359)
S. Typhimurium farm (389)
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
QRA and existing models
� 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
QRA and existing models
� 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