Sand, Small Stones

Fact sheet:
Salmonella
Version: 09/01/2015
GMP+ International B.V.
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Content
General Summary
Summary of GMP+ products standards for the animal feed sector
More Facts
1. Nature, history and prevalence of salmonella
2. Transmission to the environment, plants, animals and humans
3. Diagnose of poisoning
4. Potential hazards and adverse effects
5. Severity of the hazard
6. Standards
7. Analysis methods
8. Control measures
9. References
10. Websites
APPENDIX / APPENDICES
Fact sheet: Salmonella 2 / 57

Version: 09/01/2015 GMP+ International B.V.
General Summary

Name: Salmonella
code: M04ab
Description: A bacteria group from the Enterobacteriaceae family. They occur in the intestinal
system of animals and humans and in the environment.
Type: microbiological
Severity: high

Control measures: - (Locked) closed storage
- (Locked) closed transport
- After gathering direct transport to the client, with the advice To be fed direct or
silage / acidification'.
- Air filtration
- Analyse intermediate product(s)
- Analysis of spring/treated water quality
- Avoid extreme storage and burst bags
- Avoid grazing on pastures with chicken manure
- Bacteria strains with Biosafety-class 1 certification
- Bark peeling (or entry check)
- Blood from animals approved ante mortem
- Brewer's yeast is present in intrinsic microbiological beer
- Check of odour and appearance
- CIP (cleaning in place)
- Closed production system
- Correct storage duration and temperature
- Covering hay bales
- Covering production area.
- Cultivation conditions
- Dairy filling: carry out product and process control
- Dedicated storage/transport
- Distillation temperature min. 200C
- Dry or aerate products
- Dry plant/dry storage conditions
- Drying
- Entry and intake check: clean eggs which are not broken
- Entry and reception check
- Establish and monitor cooking temperature and time
- Establish blood purchasing requirements
- Establish purchasing requirements
- Extraction with ethanol/hexane
- Extraction with ethanol/nitric acid
- Feed beer is intrinsically microbiologically safe
- Finished product analysis
- Gathering in a clean and dry location
- Grinding line: extra attention to cleaning
- Heat treatment later in process
- Heating
- High temperature and high pressure
- Intake check: organoleptic, colour and plasma
- Low moisture percentage
- Maximum / low AW
- Maximum Aw liquid-rich end products
- Maximum pH
- Maximum pH liquid-rich end products
- Maximum temperature
- Microbiological analysis on transport air
- Monitoring cooking, drying (and decontamination) process
- Monitoring minimum temperature
- Packaged end products
- Packaged raw materials
- Pasteurisation temperature and time
- Preservation
- Prevent mouldy silage
- Prevention accumulation of organic material
- Prevention of overheating: max AW, storage duration and temperature
- Purchase animals of known origin
- Purity strains control
- Quick cooling after pelletizing
- Quick processing of the product
- Received product/raw material analysis
- Roasting: temperature and time
- Rotation of crops
- Selection of sick salmons
- Stainless steel transport/installation
- Sterilization time and temperature
- Supplier contract
- Taking measures to avert birds
- Training farmers on Good Agricultural Practices
- Use clean seawater
- Use new packaging material
- Use tapwater
- Veterinary approval
- Visual inspection transport truck
- Water management
The control measures specified in this fact sheet are all control measures which can be used depending on the
product and/or process step.

SUMMARY OF GMP+ SPECIFIC FEED SAFETY LIMITS FOR THE ANIMAL FEED SECTOR
Contaminant Product Action limit(1) Rejection limit(1) Source Supplementary Analysis method7
requirements
Microbiological: Microbiological contamination
M4a Salmonella Consumption chick feed: end products and feed GMP+ OZM Part 2; M-2a
materials for: to M2e
- Top breeding consumption chicks - 0+% 20(approaching
0%)
- Breeding increase consumption chicks - 0+% 20(approaching
0%)
- Increase consumption chicks - 0+% 20(approaching
0%)
- Consumption chicks - 0+% 20(approaching

0%)
Laying poultry feed: end products and feed GMP+

materials for:
- Top breeding laying poultry - 0+% 20(approaching
0%)
- Raising increase laying poultry - 0+% 20(approaching

0%)
- Increase laying poultry - 0+% 20(approaching
0%)
- Laying-hens and breeding hens 1% 0+% 20(approaching

0%) for S.
enteritidus and S.
typhimurium
Turkey feed: end products and feed materials for: GMP+
- Raising increase turkeys - 0+% 20(approaching

0%)
- Increase turkeys - 0+% 20(approaching

0%)
- Consumption turkeys - 0+% 20(approaching
0%)
Other animal feeds, feed materials and wet mixes - Absent in 25 gr GMP+
intended for cattle farms (except for poultry feeds).
From animal by-products derived products with the - Absent in 25 gr Commission n = 5, c = 0, m = 0, M = 0
14
exception of canned petfood Regulation (EU)
142/2011, annex
XIII, chapter 2

Contaminant Product Action limit(1) Rejection limit(1) Source Supplementary Analysis method7
requirements
From animal by-products derived canned petfood - Commission Canned petfood which
Regulation (EU) has been subjected to
142/2011, annex heat treatment with a Fc
XIII, chapter 2 value of at least 3
M4b Salmonella preservation - Feed materials for delivery to cattle farms, and Maximum pH for GMP+ If preservation can be OZM Part 2; M-5
determined through pH guarantee: achieved at a higher pH
then this should be
supported with data.
- Wet mixes for delivery to livestock farmers on the
basis of:
- Spontaneous lactic acid fermentation - 4.5 These norms do not apply

if the products are
- Add organic acids 4 supplied at a temperature
of at least 60C and the
- Add inorganic acids 3,5 supplier is demonstrably
informed of the storage
conditions.
The absence of
Salmonella can also be
shown in heat-treated wet
mixes and feed materials
(<13% moisture) through
compliance with the
norms for
Enterobacteriaceae.
[1 ] Action limit: A feasible limit agreed in consultation with the sector, supplier or customer. If this limit is exceeded then an investigation into the cause should be undertaken and corrective measures should be
taken to remove or control that cause. Maximum levels in mg/kg (ppm) of the feed materials or compound feeds, derived to a moisture content of 12% unless mentioned differently.
Rejection limit: A feasible limit agreed in consultation with the sector, supplier or customer. If this limit is exceeded then the product is not suitable for use as feed material or animal feed. Maximum levels in mg/kg
(ppm) of the feed materials or compound feeds, derived to a moisture content of 12% unless mentioned differently.
[7] The research methods (OZM) can be found via the PDV website (www.pdv.nl ; quality; research methods)
[20] Explanation of 0+: this norm does not apply to each individual sample. In a particular period of time the Salmonella incidence at company level should approach 0% (= 0+).
[14] n = number of samples to be tested; m = threshold value for the number of bacteria; the results will be considered to be satisfactory if the number of bacteria in none of the samples is greater than m, M =
maximum value for the number of bacteria; the result shall be considered unsatisfactory if the number of bacteria in one or more samples is M or more; c = number of samples for which the bacteria count gives a
result between m and M and where the sample is still considered acceptable if the result of the bacteria for the other samples is not higher than m.

More facts
Chemical name
Not applicable.
CAS-number
Not applicable.
Synonyms
Not applicable.
This fact sheet focuses on Enterobacteriaceae and Salmonella contamination of feed1 as a

possible source of contamination of food producing animals and consequently of food
products of animal origin. Other sources of Enterobacteriaceae and Salmonella contamination
will not be addressed to.
1. Nature, history and prevalence of Enterobacteriaceae and Salmonella

A. Enterobacteriaceae
The Enterobacteriaceae are a large family composed of approximately 35 bacterial genera.
The organisms are rod-shaped, Gram-negative facultative and are facultative anaerobe. Most
species grow well at 37C. There is substantial variation in the ecology, host range and
pathogenic potential of the Enterobacteriaceae. They may be found as part of the intestinal
bacterial flora of humans and other animals, others are found in water, soil and vegetables.
The term coliform refers to members of the Enterobacteriaceae which ferment lactose sugar
(EFSA13, 2007). An overview of some Enterobacteriaceae genera and species is given in table
1.
Table 1. Some Enterobacteriaceae genera and species (Stevens et al., 2003; WHO/FAO, 2008)
Genera Species
Cronobacter Enterobacter sakazakii
Eschericia Eschericia coli (E. coli)
Klebsiella Klebsiella pneumonia
Enterobacter Enterobacter amnigenus
Citrobacter Citrobacter freundii
Salmonella Salmonella enteritidis; Salmonella typhimurium
Shigella Shigella sonnei
Yersinia Yersinia pestis; Yersinia pseudotuberculosis
The determination of Enterobacteriaceae to assess the hygiene of production and the quality
of food and feed has been introduced and found to be useful as indicators of hygiene and
contamination (Schothorst and Oosterom, 1984; Gilbert et al., 2000). Enterobacteriaceae are
used as process-indicators because they die at 60C and above and are an indication for the
effectiveness of heat treatment (Ridderbos, 2006). In feed this has been demonstrated by
Stott et al. (1975). There is a recognised association between the risk of isolation of
Salmonella and Sakazakii and degree of Enterobacteriaceae contamination (Veldman et al.
1995; EFSA14, 2004; EFSA13, 2007). An association between Salmonella and
Enterobacteriaceae has also been found by several authors (Veldman et al., 1995), however
in the EFSA13 (2007) report the association between Salmonella and Enterobacteriaceae in
infant formula was not found. In the Netherlands Enterbacteriaceae are included in hygiene
guidelines of various food industries as a hygiene indicator (nVWA, 2011).
1
Any substance or product, including additives, whether processed, partially processed or unprocessed, intended to be used for
oral feeding to animals

Enterobacteriaceae are also as an indicator of feed hygiene. Schothorst and Oosterrom
(1984) suggested the use of Enterobacteriaceae counts (EC) for assessment of good
manufacturing practices in rendering plants. Stott et al. (1975) demonstrated a correlation
between EC and Salmonella contamination in pelleted feeds. Cox et al. (1983) reported no
correlation between EC and Salmonella. Veldman et al. (1995) found that Enterobacteriaceae
were useful markers of both the Salmonella contamination level and the efficiency of
decontamination by pelleting.
There are Enterobacteriaceae that are enteroinvasive and that are enterotoxigenic, causing
two major types of infection: intestinal and extra-intestinal. Many of the organisms are
commensal members of the intestinal microbial flora, whilst others can cause intestinal and
invasive diseases. A number can cause diarrhoeal diseases, e.g. most Salmonella serovars
and Shigella dysenteriae, whereas many others are opportunistic pathogens which may not
only cause intestinal disease, but also bacteraemia, meningitis, urinary tract, respiratory and
wound infection (EFSA13, 2007).
In the following chapters Enterobacteriaceae will not be addressed specifically because of the
numerous genera and species present within this group and the Enterobacteriaceae
information might be to general and incomplete concerning the specific bacteria.
B. Salmonella
Salmonella is a gram-negative, rod-shaped non-spore-forming genus of the family
Enterobacteriaceae (RIVM2, 2006). It is named after dr. Salmon who, together with his
assistant, discovered Salmonella (USDA, 2011). The taxonomy and nomenclature of
Salmonella have changed over the years and are still evolving. The nomenclature in use at the
CDC (Centers for Disease Control and Prevention) is shown in table 2.
Table 2. CDC Salmonella nomenclature (Brenner et al., 2000).

Family Genus Species No. Serogroups Serotypes
Subspecies
Enterobacteriaceae Salmonella S. enterica 6 50 >2300
S. bongori 1
More than 2500 serotypes exist and the prevalence of the different serotypes changes over
time (EFSA1, 2009). Two serotypes, Salmonella enteritidis (S.e.) and Salmonella Typhimurium
(S.t.) are the most common nowadays (EFSA, 2009; USDA, 2011; RIVM2, 2011). Several
harmonised baseline surveys have been conducted in different populations of food production
animals and this has procured information on serotype distributions as was presented by the
EFSA1 (2009) and is demonstrated in table 3.
Table 3. Distribution of Salmonella serotypes in human isolates and isolates from EU baseline surveys in broilers
1
layers, turkeys and slaughter pigs (EFSA , 2009).
Salmonella Humans Broilers Laying hen Turkeys Slaughter pigs
serotype (N=138707) (N=1448) flocks fattening flocks (N=2600)
(N=1486) (N=1084)
S. Enteritidis 82251 538 899 55 126
S. Typhimurium 21136 65 123 86 1040
S. Infantis 1331 295 171 72 49
S. Virchow 1106 30 41 11 7
S. Newport 771 8 11 33 24
S. Stanley 669 0 0 0 0
S. Hadar 488 59 53 152 8
S. Derby 475 13 14 123 380
S. Kentucky 435 44 12 1 0
S. Agona 421 16 38 31 28
Other important serotypes in Salmonella positive tested isolates, not mentioned in table 2, are
S. Mbandaka (broiler and laying hen) (EFSA2,3, 2007); S. Livingstone (laying hen) (EFSA3,

2007); S. Bredeney, S. Saintpaul , S. Kottbus, S. Orion and S. Blockley (turkey) (EFSA4,
2008); S. Rissen and S. 4,[5]12:i:- (slaughter pigs) (EFSA5, 2008).
In Appendix II an overview is given of the trends in Salmonella serotypes most important in
humans.
A microscopic picture of Salmonella is shown in figure 1.
Figure 1. Salmonella (red colour) (source: Wikipedia)
Depending on the serotype the effect of different environmental circumstances of Salmonella

varies. Important environmental parameters that affect the activity of Salmonella are:
temperature, pH and water activity (aw) (Giaouris et al., 2005). In general Salmonella is
relatively sensitive to heat and cold, but there are exceptions (Henry et al., 1969; Garibaldi et
al., 1969; Srqvist, 2003; Food-info, 2012). The water activity affects the heat resistance
(Goepfert et al., 1970; Mattick et al., 2000). Salmonella ordinarily is also sensitive to high
acidity (Keng Chee Chung and Goepfert, 1970; US-FDA1, 2009).
Salmonella has long been recognised as an important zoonotic pathogen of economic

significance in animals and humans (EFSA1, 2009).
Although opinions vary on the importance of feed contamination there is evidence that feed
can pose a risk of Salmonella infection for humans and animals. Transmission of Salmonella
from feed to animals consuming the feed, and to food products derived from the animals has
been shown (EFSA6, 2008). The overlap between Salmonella serotypes commonly found in
feed and those isolated from human cases of salmonellosis is limited, but across the EU, four
of the serotypes ranked in the top ten feed isolates (Infantis, Typhimurium, Agona and
Enteritidis) are also in the top ten public health serotypes (EFSA7, 2006). sterberg et al.,
(2010) however point out that the serotypes commonly detected in feed often differ from the
serotypes most often isolated from animals and humans. Hald et al. (2006) estimated that
2.1% of domestically acquired infections in the period 1999-2003 could be attributed to feed
borne serotypes acquired through the consumption of pork and beef. They concluded that
more than 90% of serotypes have the potential, if they occur in feed, for infecting humans via
production animals or foods of animal origin.
The common reservoir of Salmonella is the intestinal tract of animals, which result in a variety
of foodstuffs covering both food of animal and plant origin as sources of infections.
Transmission often occurs when organisms are introduced in food preparation areas and are
allowed to multiply in food, e.g. due to inadequate storage temperatures, inadequate cooking
or cross contamination of ready-to-eat food. The organism may also be transmitted through
direct contact with infected animals or humans or faecally contaminated environments (EFSA1,
2009).
The widespread and emerging antimicrobial resistance of certain Salmonella serotypes
threatens to increase treatment failure, relapse and death due to e.g. enteric fever (WHO1,
2005; Gupta et al., 2008) is of great concern.

2. Transmission and likelihood of occurrence
Environment
Salmonella grows between 7C and 49.5C but it grows best between 35C and 37. It grows
through a pH range of 3.8 to 9.5, has a minimum water activity level of 0.94 and grows in
either the presence or absence of oxygen (CCC, 2012).
Salmonella bacteria are readily killed by heating to 60C for 2 to 6 minutes or 70C for 1
minute (CCC, 2012). However some serotypes are known to be heat-resistant, e.g. S.
senftenberg 775W (Henry et al., 1969; Goepfert et al., 1970; Maas et al., 2003). Heat
resistance increases with reducing aw-values (Goepfert et al., 1970; Mattick et al., 2000).
Salmonella can survive for long periods in low water content foods. Salmonella can also
survive for long periods in food under refrigeration, up to 28 days on the surfaces of
vegetables. Death occurs during the freezing process, but those that survive remain viable
during frozen storage. Salmonella survives well in foods and on surfaces. Survival in dry
environments is a characteristic of Salmonella (CCC, 2012).
Manure is a possible source of Salmonella contamination of soil (Natvig et al. 2002; Islam1,2
et al., 2004). When manures are applied to land, there is likely to be some movement of
microorganisms through the soil matrix, both vertically and horizontally. Factors known to
influence the horizontal movement of microorganisms across soils include soil type, soil water
content, amount and intensity of rainfall, temperature, nematodal activity, surface charge,
transport through plant roots, and soil pH. Generally, the microorganisms survival is favoured
in aqueous environments, and thus water availability and movement are the single most
important factors in determining how far pathogens are likely to move through or across soils.
Several authors studied the survival of Salmonella in soil. Repeated freeze-thaw cycles reduce
the level of S.t. in soils but may not completely eliminate this organism (Natvig et al., 2002).
Barak and Liang (2008) showed that S. enterica could survive for up to six weeks in fallow soil
(ploughed an harrowed but left unsown for a period) with the ability to contaminate crop.
Manure compost has also been shown to play a role in contaminating soil and root
vegetables with Salmonella, up to several months (Islam1,2 et al., 2004).
Salmonella may be found in water sources. These water sources have been in contact with
Salmonella contaminated faeces of infected humans or manure of infected animals.
Additionally contaminated waste can enter the water through different ways, including sewage
overflows, sewage systems that are not working properly, polluted storm water runoff, and
agricultural runoff. Wells may be more vulnerable to such contamination after flooding,
particularly if the wells are shallow, have been dug or bored, or have been submerged by
floodwater for long periods of time (CDC, 2009).
Another water source of risk is irrigation water. Contaminated irrigation water can
contaminate soil and consequently contaminate root vegetables with Salmonella for several
months (Islam1,2 et al., 2004).
Water in water/tank vessels, used as drinking water for animal, can also be contaminated
with Salmonella (Taylor et al., 2000; Teplitski, 2009). In the case of a waterborne outbreak of
S. Saintpaul, described by Taylor et al. (2000), frogs and/or mice may have been the original
source of contamination. Once pathogens contaminate water vessels, they become
established and form biofilms (bacterial communities encased in a protective slime layer)
(Teplitski, 2009).
Pests (e.g. rodents, birds) are a possible source of Salmonella. They can amplify the number
of pathogens in the environment and transfer them. Wild birds and mammals are generally
regarded as the main reservoir for Salmonella (Meerburg and Kijlstra, 2007). Within poultry
farms, infected rodents are often reported. Henzler and Opitz (1992) found a high prevalence
(24%) of S.e. in rodents present on S.e. contaminated chicken layer farms. S.e. was not
detected in mice on clean farms. This is confirmed by a study of Garber et al. (2002) who

found that the presence of S.e. in layer houses was associated with, amongst others, number
of rodents trapped. The prevalence of S.e. in mice from environmentally positive houses was
nearly four times that of mice from environmentally negative houses. Besides pests, other wild
animals are also known to carry Salmonella (e.g. badgers (Wilson et al., 2003 and foxes
(Kahn, 2005)) or domesticated animals (e.g. dogs (Morse and Duncan, 1975; Joffe and
Schlesinger, 2002) and cats (Van Immerseel1 et al., 2004)).
Insects are also known vectors of Salmonella (Bates et al., 2004). Several Salmonella
serotypes have been isolated from insects e.g. flies (Bailey et al., 2001 Holt et al., 2007).
These insects frequent fecal deposits where they could encounter salmonellae and may
transport these bacteria to foods, feed and drinking water for farm animals (Doyle et al., 2009).
Salmonella can also be spread via the atmosphere. Atmospheric spreading can occur via
fan-driven air, dust, fluff/down or water droplets (Berrang et al., 1995; Mitchell et al., 2002).
Airborne movement of dust and fluff has been implicated in the transfer of this organism in
animal husbandry (Mitchell et al., 1995; Chinivasagam et al., 2009). In feed mills dust, besides
feed ingredients, has also been indicated as a major source of Salmonella contamination
(Jones and Richardson, 2004). More specifically Torres et al. (2011) demonstrated that dust of
feed mill intake pits have an increased risk of being contaminated with Salmonella. Feed mills
can be endemically contaminated with Salmonella as was demonstrated by Davies and Wales
(2010). The endemic contamination was reflected in isolates obtained from finished products
and destination flocks. Renovation of equipment and chemical treatment of equipment and
feed had not removed endemic strains,
Plants
General sources of Salmonella infection:
In general any products of vegetal origin can be contaminated with Salmonella via manure
(WHO1, 2005), manure compost (Islam1 et al., 2004), water (e.g. irrigation water) (Islam1,2 et
al., 2004; Lapidot and Yaron, 2009), pests (e.g. rodents, birds) (Meerburg and Kijlstra, 2007)
or via the atmosphere (Torres et al., 2011). The contamination of products of vegetal origin
via adherent soil originating from manure is also reported (Natvig et al., 2002). Barak and
Liang (2008) studied the role of Salmonella contaminated soil, the potential for crop debris to
act as inoculum from one crop to the next. They found that subsequent crop could be
contaminated via contaminated crop debris originating from contaminated soil.
Other sources than feed ingredients of Salmonella contamination of animal feed:

Besides the products themselves another important factor influencing Salmonella
contamination / growth are processing and storage conditions. Examples are improper
heating and temperature abuse during processing and storage (Juneja et al., 2009).
The EFSA6 (2008) report that feed ingredients can be also contaminated during processing
because of a residual contamination of the premises, equipment and staff. This risk of
contamination can be increased because of an insufficient cleaning and disinfection of the
premises and equipment used to process the feed ingredient, but also because of poor
hygiene conditions during storage, poor hygiene of the staff, or environmental
conditions (moisture and temperature) favourable for Salmonella multiplication. Salmonella
can also be spread through operators, vermin, cross contamination, recontamination
after processing, etc. (EFSA6, 2008).
Concerning oil seed meals, the flow of air used in the meal dryer cooler (DC) and pellet
cooler enables re-entry of airborne Salmonella. Potential conditions exist in the DC and pellet
cooler for water to condense and form a wet, warm environment, ideal for Salmonella to
rapidly multiply. So the most prevalent measure point of Salmonella contamination is the dust
from the DC and pellet cooler cyclone collectors (Kemper, 2010).
The conditions of transport and storage of feed ingredients also affect the risk of Salmonella
contamination. The risk is increased by poor hygiene and no respect of good practices, mainly
poor cleaning and disinfection of containers, wheels, equipment for collection, and
silos, and also by the presence of vermin and wild birds (EFSA6, 2008).

In the section Animals Exposure other routes of Salmonella contamination will be
discussed. This information might also apply to vegetal feed ingredients.
The contamination risks present at the feed mill are mainly if the ingredients are contaminated
with Salmonella upon arrival at the feed mill or the feed mill environment or the feed mill
environment carries a permanent Salmonella infection that will contaminate the ingredient
(EFSA6, 2008) or animal feed. Experience has shown that a feed mill that received a
contaminated ingredient may become contaminated for an extended period of time (EFSA6,
2008).
Unloading ingredients in the intake pit creates large amounts of dust, which may carry
Salmonella infection to the premises (EFSA6, 2008). Torres et al. (2011) reports that intake
pits were demonstrated to have an increased risk of Salmonella culture-positive dust samples.
Wierup and Hggblom (2010) also identified the intake pit / bottom part of the elevator of feed
ingredients as a critical point concerning Salmonella. The transport equipment used for
ingredients may become contaminated as well as the flat storage areas and silos due to
contaminated dust particles remaining inside the systems. If intake pits or other parts of the
transport or storage systems carry infection since the previous ingredient Salmonella negative
ingredients may also become contaminated. An efficient dust control in the pit area is very
important to prevent further spread of Salmonella from potentially contaminated ingredients.
Intake pits also easily attract vermin and wild birds and automatic doors surrounding the pit
offer some protection to contamination from faecal material from wild animals. Flat stores are
specifically attractive to wild birds and rodents and for that reason effective control measures
are important to prevent contamination (EFSA6, 2008).
Water from trucks or rail cars entering the intake pit due to rainfall or leakage through roofs
of storage buildings is not unusual and can lead to increased moisture content in the
commodity giving rise to actual multiplication of Salmonella in hot spots of the ingredient. In
feed mills situated close to the sea the lower end of elevators are usually below see level, a
situation which temporarily might generate moisture levels in the dust above the level where
Salmonella multiplication may occur (EFSA6, 2008).
Ingredients entering the feed mill with elevated temperatures or when warm days are
followed by cold nights may cause condensation of free water on cold surfaces in the
transportation systems and also in storage containers. Condensation within the feed mill
will also occur if the temperature difference is more than 5C between the pelleted feed and
the environment. It is not unusual that coolers are not efficient enough to produce a low
temperature of the pellets thus the warm pellets will give rise to condensation and free water
in clean side of the feed mill. Condensation usually occurs as droplets of water and if the
conditions favour growth, a single droplet typically in the top of the conveyer or silo, may
contain large numbers of bacteria (EFSA6, 2008).
Concerning processing, the build up of feed within the equipment increases the risk of
Salmonella contamination of feed, as is equipment being difficult to inspect and clean.
Leakages, spillages or dust accumulated on floors or elsewhere in the premises may cause
dissemination of Salmonella in the mill particularly if the leakages occur from the ingredients in
areas of the feed mill where the finished product is exposed to the environment (EFSA6,
2008). Besides the intake pit (see above) Wierup and Hggblom (2010) have identified four
other critical points in the feed mill, being:
a) Dust from the aspiration system (filter);
b) Top of pellet cooler;
c) Room for pellet coolers;
d) Top of bin for feed (compound feed).

Because of the very large amounts of cooling air used in the coolers the microbial quality of
the air is very important. Contaminated dust particles in the air will be picked up by the feed
in the cooler and potentially contaminate the cooler especially if condensation occurs in the
top (EFSA6, 2008). Other risks may be a general aspiration system for ingredients and
heat treated products. The storage bins for the aspiration dust may harbour Salmonella for
extended periods of time. Dust from the aspiration may only be reintroduced in the process
before the heat treatment (EFSA6, 2008). Torres et al. (2011) found that 12.5% of the
analysed dust samples from 523 Spanish feed mills were Salmonella positive.
The use of high temperatures to accomplish pasteurisation during processing is based on the
destructive effects of time and temperature on Salmonella and other microorganisms.
Microbiologists have identified at least 11 factors or parameters of microorganisms and their
environment that can affect heat destruction. These factors include moisture or water activity,
fat levels, presence of salts, presence of carbohydrates, pH, protein content, number of
organisms, age of organisms, inhibitory compounds, and time and temperature history.
Despite the influence these factors can have on the resistance of microorganisms to heat,
thermal destruction during the processing steps of pelleting or extrusion is the most critical
control step for destruction or reduction of Salmonella and other pathogenic microorganisms
(AFIA, 2010).
For more detailed information concerning specific feed and food of vegetal origin of risk and
Salmonella outbreaks in the EU, see Appendix III.
In the DOS database data can be found concerning Salmonella in feed of vegetal origin.
Animals
Exposure:
Animals are exposed to Salmonella via varies sources: e.g. water, feed, litter / manure, farm
staff and the environment (outdoor and indoor). Hatcheries are also possible sources of
Salmonella (WHO3, 2009).
In figure 2 possible transmission routes of Salmonella on farms are shown.

Figure 2. Possible Salmonella transmission routes on farm (modified from Veldkamp and Bokma-Bakker, 2004).
Concerning feed contamination by Salmonella is not uncommon even in feed that has
undergone heat treatment (Hacking et al., 1978; Cox et al., 1983; Jones and Richardson,
2004). EU member states (MS) data from 2005 show a national prevalence for compounded
poultry feed of 6% in one MS, while most other countries have prevalences in the range from
0% to 1.5% (EFSA7, 2006). Similar contamination rates were reported for pig (up to 1.7%) and
cattle (up to 2.4%) feed in the EU. The industry based data from 2001 and 2010, as shown in
table 4 (PDV1, 2003; PDV2, 2004; PDV3, 2005; PDV4, 2006; PDV5, 2007; PDV6, 2008; for
2008-2011 data in the DOS-database of GMP+ International were used) reports an incidence
between 0 and 1.0% of Salmonella contaminated samples in compounded feed to different
food animal species (poultry, swine and cattle). The lowest prevalence was found in feed for
top breeding poultry flocks and the highest for laying hens.
1
Table 4. Industry Salmonella data compound feed GMP+ certified (GMP+ FSA scheme) companies (PDV , 2003;
2 3 4 5 6
PDV , 2004; PDV , 2005; PDV , 2006; PDV , 2007; PDV , 2008; for 2008-2011 data in the DOS-database of GMP+
International were used).
Feed 2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011
% % % % % % % % % % %
Poultry
2
Top breeding 0 0 0 0 0.1 0 0.2 0.7 0 0 0
Raising 0.6 0 0 0 0 0 0 0 0 0 0
increase
Breeding 0.5 0.3 0.5 0.5 0.1 0.1 0 0 0 0 0.5
Broilers 0.2 0.1 0.4 0.3 0.2 0.1 0.4 0 0.1 0 0
Laying and 0.9 0.5 0.4 1 0.8 0.7 0.5 0.5 0.3 0.3 0.4
breeding hens
2
Meat and egg.

Feed 2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011
% % % % % % % % % % %
Raising 0 0 0 0 0 0 0 0 0 0 0
breeding
turkeys
Breeding 0 0 0 0 0 0 0 0 - - -
turkeys
Meat turkeys 0 0 0 0.7 0 0 0.8 1.6 0 0 0
Cattle 0 0.9 0.7 0.4 0.5 0.3 0.2 0.5 0.1 0.2 0.3
Horse - - - - - - - 0 0 0 0
Goat - - - - - - - 0 0 0 0
Sheep - - - - - - - 0 0 0 0
Swine 0.3 0.6 0.6 0.6 0.4 0.3 0.2 0.5 0.2 0.3 0.2
The EFSA summarises EU MSs Salmonella data in compound feed in annual EU summary
reports, as shown in table 5. The range within the MSs is also reported and shows a wide
range within the MSs. However due to significant differences in monitoring and reporting,
strategy data are not comparable between MSs, and cannot be considered as national
prevalence (EFSA9, 2010). The reported percentages of positive single samples/batches might
not always be representative of feed on the national markets, as it might reflect intensive
sampling of high-risk products (EFSA10, 2011). For more information concerning these data is
referred to the cited EFSA references.
9 10 12
Table 5. Salmonella (%) in compound feed (EFSA , 2010; EFSA , 2011; EFSA , 2012).
Compound 2006 2007 2008 2009 2010
feed (range) (range) range) (range) (range)
Cattle 0.7 (0-9.4) 0.5 (0-8.0) 0.5 (0-3.6) 0.4 (0-4.9) 0.7 (0-9.1)
Pig 0.6 (0-3.3) 0.8 (0-5.9) 0.6 (0-3.6) 0.7 (0-2.9) 0.5 (0-3.6)
Poultry 0.6 (0-3.7) 1.0 (0-10.1) 0.9 (0-8.3) 1.0 (0-18.0) 0.5 (0-1.6)
Salmonella serotypes data in animal feeds monitored by the U.S. Food and Drug
Administration (FDA) (2002-2009) are shown in table 6. The 25 most common Salmonella
serotypes found in different categories of animal feeds are shown.

Table 6. 25 most common Salmonella serotypes found in different categories of animal feeds (2002-2009) in the
U.S. (Li et al., 2012).
Rank Serotype %
1 Senftenberg 8.9
2 Montevideo 8.9
3 Mbandaka 8.6
4 Tennessee 6.2
5 Typhimurium 5.4
6 I 4, [5], 1 2:i;- 5.0
7 Schwarzengrund 4.7
8 Anatum 4.3
9 Agona 3.5
10 Johannesburg 3.5
11 Enteriditis 3.1
12 Havanna 3.1
13 Cerro 2.7
14 Oranienburg 2.7
15 Arkansas 1.6
16 Bredeney 1.6
17 Cubana 1.6
18 Derby 1.6
19 Alachua 1.2
20 Hadar 0.8
21 Weltevreden 0.8
22 Amager 0.8
23 Muenchen 0.8
24 Kentucky 0.8
25 Lille 0.4
All other serotypes 17.5
A major Salmonella Thompson outbreak occurred in the Netherlands in 2012. The cause was
traced back to smoked salmon of one producer. The serotype has, up to the moment of
publishing this fact sheet, not been found in Salmonella contaminated animal feeds as present
in the DOS database of GMP+ International. Salmonella Thompson was also not reported in
the top 25 serotypes found in animal feed in the U.S. between 2002 and 2009 (Li et al., 2012).
For more information concerning other sources than feed ingredients of Salmonella
contamination of animal feed, see Chapter 2, section Plants - Other sources than feed
ingredients of Salmonella contamination of animal feed.
For more detailed information concerning specific feed and food of vegetal and animal origin
of risk and Salmonella outbreaks in the EU, see Appendices III and IV.
Several authors have studied the occurrence of Salmonella infections in free-range animals
and compared organic and conventional farming systems. The results of these studies are
conflicting. The results of the study of Alali et al. (2010) suggest that the prevalence of fecal
Salmonella was lower in organic (free-range) birds than in conventionally raised birds.
Conversely Kijlstra and Eijck (2006) state that health problems in organic livestock farming are
often related to the outdoor access area, exposing the animals to various bacterial infections.
Bailey and Cosby (2005) specifically address Salmonella and state that free-range chickens
have access to the outside, where there is sufficient opportunity for exposure to wild birds,
insects, rodent droppings, and other potential carriers of Salmonella. McCrea et al. (2006)
found in their study that free-range chickens had the highest prevalence of Salmonella, in
comparison with other poultry.
In the DOS database data can be found concerning Salmonella in feed.

Absorption:
Gastric acidity is a major barrier to colonization and infection of the gastrointestinal tract by
microorganisms, e.g. Salmonella, and is a first line of defense against microbial pathogens
that infect their host via the oral route (Bearson et al., 2006). The EFSA6 (2008) assumes that
Salmonella can pass the stomach due to a rapid passage and thus, too little acidification is
plausible. Contamination of feed usually results in intestinal colonisation, and a long term
shedding of Salmonella. As is confirmed by Hohmann et al. (1978) who found that within 3
hours after oral challenge of mice with S.t., foci of infection developed in the Peyers patches
of the small intestine. Barrow et al. (1988) studied the intestinal Salmonella colonisation in
three-week-old chickens and found that Salmonella persisted longest in the caeca and were
also present for a shorter time in the crop. Concerning ruminants Salmonella is also found in
the rumen (Brownlie and Grau, 1967; Mattila et al., 1988). McEvoy et al. (2003) found that 2%
of the rumen samples collected from an Irish beef abattoir were contaminated with Salmonella.
It is documented that, when animals are starved, Salmonella can survive and multiply in the
rumen (Brownlie and Grau, 1967; Teklu and Negussie, 2011).
Distribution:
Sivula et al. (2008) found that in one-week-old chickens and in mice, besides ceacal
colonisation, systemic organs are most highly colonised as well. In mice, systemic colonisation
increased with time after infection. This slowly increasing systemic colonization might be due
to continuous re-seeding of systemic organs from highly colonized areas of the gut, including
the cecum, via gut associated lymphoid tissues. The spleen and liver were increasingly
colonised in mice throughout the infection. Teirlynck et al. (2009) also found Salmonella
colonisation in the spleen and liver of broilers. Salmonella also colonises in the tissues of the
chicken ovary and oviduct, consequently contaminating eggs. Data of Keller et al., (1995)
show that prior to egg shell deposition, forming eggs are subject to descending infections from
colonized ovarian tissue, ascending infections from colonized vaginal and cloacal tissues, and
lateral infections from colonized upper oviduct tissues.
Chattopadhyay et al. (2010) studied the effect of Salmonella infection in pregnant mice and
found massive placental infection and Salmonella was localised in the deeper layers of the
placenta. Pejcic-Karapetrovic et al. (2007) demonstrated that rapid proliferation of Salmonella
in the placental environment of pregnant mice. Two hours after infection, only about 5% of the
infective dose reached the placenta, yet this number grew astoundingly by 14 hours. In
contrast, two hours after infection relatively higher numbers homed to the spleen, but they
expanded only marginally in the first 24 h. This marked contrast in the ability of Salmonella to
proliferate in the spleen versus placenta is suggestive of a unique escape and/or invasive
mechanism in the pregnant host.
Salmonella has also been isolated from the udder of cows (Wood et al., 1991).
Metabolism:
Not applicable.
Excretion:
Salmonella infected animal are able to shed Salmonella into the environment for extended
periods of time increasing the risk for environmental contamination, spread of the organism
within the flock or herd and contamination of the food supply (Sivula et al., 2008). Faecal
shedding was demonstrated in poultry (Barrow et al., 1988; Barrow et al., 2004), swine (Molla
et al., 2010) and ruminants (Brownlie and Grau, 1967). McEvoy et al. (2003) found that 2% of
the faecal samples collected from an Irish beef abattoir were contaminated with Salmonella.
Wood et al. (1991) found that the milk of a Salmonella infected cow, with an infected quarter of
the udder, was also contaminated with Salmonella. The means by which the organism gained
entrance to the udder of this animal was unknown. El-Ziney and Al-Turki (2007) state that milk
might get contaminated either by faecal contamination or by direct excretion from the udder
into milk. However no scientific data were found to support this statement.

Humans via animal products
Although opinions vary on the importance of feed contamination there is evidence that feed
can pose a risk of Salmonella infection for humans and animals (EFSA6, 2008). However the
hygienic conditions during the slaughter process and processing of products of animal origin
are of critical importance in controlling Salmonella contamination (WHO4, 2002; Botteldoorn et
al., 2003; Delhalle et al., 2008).
The WHO2 (2012) reports that Salmonella is generally transmitted to humans through the
consumption of Salmonella contaminated food of animal origin, mainly meat, poultry, eggs
and milk. The EFSA1 (2009) concludes in the community summary report on trends and
sources of zoonoses and zoonotic agents in the European Union in 2007 that overall, reported
data from 2007 underline the generally accepted conclusion that the main sources of
Salmonella infections in humans are from different types of meat (pig, poultry and bovine) and
eggs. Salmonella was most often reported in fresh meat and products of meat origin,
particularly in poultry meat followed by pig meat. In the other food categories, Salmonella was
found less frequently: occasionally from table eggs, fishery products, but seldom from milk
and cheeses (EFSA1, 2009). The RIVM2,3 (2011; 2012) reports that in the Netherlands, as in
other EU MSs, eggs have been the most important source for human salmonellosis for the
past 20 years.
For more information concerning other sources than feed ingredients of Salmonella
contamination of animal feed, see Chapter 2, section Plants - Other sources than feed
ingredients of Salmonella contamination of animal feed.
For more detailed information concerning specific feed and food of animal origin of risk, see
Appendix IV.
In the DOS database data can be found concerning Salmonella in feed of animal origin.
Humans
Exposure:
In 2010, the number of salmonellosis cases in humans decreased by 8.8 % compared with
2009, and the statistically significant decreasing trend in the European Union continued for the
sixth consecutive year. In total, 99,020 confirmed human cases were reported in 2010. It is
assumed that the observed reduction in salmonellosis cases is mainly due to successful
Salmonella control programmes in fowl populations (EFSA12, 2012). In 2007 155540 confirmed
cases of human salmonellosis were reported from 30 countries, including 27 EU MS sand
three non-MSs (EFSA1, 2009).
The EFSA concludes in the community summary report on trends and sources of zoonoses
and zoonotic agents in the European Union in 2007 that overall, reported data from 2007
underline the generally accepted conclusion that the main sources of Salmonella infections in
humans are from different types of meat (pig, poultry and bovine) and eggs. Salmonella was
most often reported in fresh meat and products of meat origin, particularly in poultry meat
followed by pig meat. In the other food categories, Salmonella was found less frequently:
occasionally from table eggs, fishery products, but seldom from milk and cheeses (EFSA1,
2009). The WHO1 (2005) also report that salmonellosis in humans is generally contracted
through the consumption of Salmonella contaminated food of animal origin (mainly meat,
poultry, eggs and milk), although many other foods, including green vegetables contaminated
from manure, have been implicated in its transmission.
Salmonella serotypes Enteritidis and Typhimurium represent 70% of the analysed isolates in
the Netherlands (RIVM2, 2011).

Absorption:
In humans Salmonella invades and penetrates the epithelial surfaces of the intestine
(McCormick et al., 1983; Elsinghorst et al., 1989) by translocation across specialized Peyer's
patch epithelium and then proliferating in the mucosal macrophages (Kraus et al., 1999).
Distribution:
Salmonella-bound to lymphocytes have been detected in the peripheral blood of typhoid
patients (Jirillo and Antonaci, 1985) and it is likely that these complexes
Salmonella/lymphocytes could be ingested by splenic macrophages (Altamura et al., 2001).
Nix et al. (2007) confirm that Salmonella can colonise the human spleen and also liver and
mesenteric lymph nodes.
Trans-placental transfer of Salmonella has been demonstrated in humans (Zettel et al., 1995).
Metabolism:
Not applicable.
Excretion:
Salmonella is excreted via breast milk (Qutaishat et al., 2003; Chen et al. 2005; Cooke et al.,
2009), urine (Qutaishat et al., 2003) and faeces (Cooke et al., 2009).
3. Diagnose of poisoning
Animals
Clinical diagnosis is supported by isolation of salmonellae. Preferred tissues for sampling
include ileum, ileocecal lymph nodes, tonsil, and cecum. In live animals, tonsil scrapings are
preferable to rectal swabs for isolation because of the unpredictability of fecal shedding in
asymptomatic carriers. Isolation alone is not sufficient for a definitive diagnosis due to the
ubiquitous nature of the organism in clinically normal animals. Bacteriology results should be
supported by histology and other tests, such as phage typing or polymerase chain reaction.
Current serological tests are neither sensitive nor specific enough to be used for individual
animal diagnosis, but are essential in determining the prevalence of asymptomatic carrier
animals and are thus essential to herd control measures (Alsop, 2005). In the Merck
Veterinary manual (Kahn, 2005) the faeces of wild rodents and birds that ma in habit the
premises are also mentioned useful to be tested. Concerning faeces of the food-producing
animals culture techniques that involve suppression of faecal E coli are usually necessary, and
several daily faecal cultures may be necessary to isolate the organism. Blood cultures in
septicemic animals may be used. Serologic testing is difficult to interpret (Kanh, 2005).
Also history of exposure and analysis of feed and water can be included.
Humans
Chaicumpa et al. (1992) isolated S. typhi in blood, stool and bone marrow of patients with
clinical typhoid fever. They also used an enzyme-linked immunosorbent assay (ELISA) for
detecting S. typhi antigens in urine and concluded that since S.typhi antigen is intermittently
excreted in the urine of patients with typhoid fever, serially collected urine from patients with
typhoid should be tested for antigen 9. In the Merck Manual the use of antigens in diagnosing
is also mentioned. Typhoid bacilli contain antigens (O and H) that stimulate the host to form
corresponding antibodies. A four-fold rise in O and H antibody titers in paired specimens
obtained 2 weeks apart suggests S. typhi infection (Beers et al. 2006).
In the Merck Manual is stated that cultures of blood, stool and urine should be obtained. Blood
cultures are usually positive only during 2 weeks of illness, but stool cultures are usually
positive during the 3rd to 5th week. If these cultures are negative and typhoid fever is strongly
suspected, culture from a bone marrow biopsy specimen may reveal the organism (Beers et
al., 2006).

Concerning non-typhoidal Salmonella diagnosis is by isolating the organism from stool or
another infected site. In bacteremic and focal forms, blood cultures are positive, but stool
cultures are generally negative (Beers et al., 2006).
Also history of exposure and analysis of food and water can be included.
4. Potential adverse effects
Environment
No data were studied.
Animals
Young animals have a greater susceptibility for Salmonella that may be due to the high
gastric pH, absence of a stable intestinal flora and limited immunity (Kahn, 2005).
In animals, Salmonella infections cause two separate problems: clinical salmonellosis and a
clinically silent carrier state. Inapparent long-term carrier animals may shed salmonellae in
faeces either intermittently or continuously. The carrier state is much more common than
clinical disease (Alsop, 2005; EFSA6, 2008). Salmonella may than easily spread between
animals in a herd or flock without detection and other animals may become intermittent or
persistent carriers (EFSA9, 2010).
Salmonella infection of food-producing animals has differing manifestations according to the
livestock species and Salmonella serotype(s) involved (EFSA6, 2008). The evidence of
surveillance in Europe and elsewhere (Davies2 et al., 2004; EFSA7, 2006; EFSA11, 2007;
EFSA10, 2011) is that infections of pigs and poultry are often widespread in many EU Member
States but typically asymptomatic, whilst ruminants may be less frequently infected but more
often show clinical signs, also in adult animals (EFSA6, 2008).
In general salmonellosis is characterized clinically by one ore more of three major syndromes:
septicemia, acute enteritis and chronic enteritis. Young calves, piglets, lambs and foals usually
develop the septicemic form. Adult cattle, sheep and horsed commonly develop acute enteritis
and chronic enteritis may develop in growing pigs and occasionally in cattle (Kahn, 2005).
Septicemia: the illness is acute. In young animals fever (40.5-41.5C) is usual and death
occurs in 24 to 48 hours (Kahn, 2005; EFSA9, 2010). In pigs, as dark red to purple
discoloration of the skin is common, especially of the ears and ventral abdomen (Kahn, 2005).
In poultry fowl typhoid (FT) and pullorum disease (PD) are the septicaemic diseases. Clinical
signs include anorexia, diarrhea, dehydration, weakness and high mortality. In mature fowl, FT
and PD are also manifested by decreased egg production, fertility and hatchability. Gross and
microscopic lesions due to FT and PD in chickens include inflammation of e.g. the liver,
spleen, caecum, heart muscle, ventricles of the brain, lung and eye. In mature fowl, lesions
include inflammation of e.g. ovaries, fallopian tubes and testes (Shivaprasad, 2000).
Acute enteritis: initially, there is fever (40.5-41.5C), flowed by severe watery diarrhea. Milk
production often declines in dairy cows. Abdominal pain is common and severe in horses.
Affected horses are severely dehydrated and may die within 24 hors of the onset of diarrhea
(Kahn, 2005). Paratyphoid infection is an acute and chronic infection of poultry and mammals.
Drooping wings, shivering and huddling near a heat source, muscular incoordination and
trembling and enteritis are signs in poultry (FAO1, 1994).
Sub-acute enteritis: may develop on farms where the disease in endemic. The sings include
mild fever (39-40C), soft faeces, inappetence and some dehydration. There may be a high
incidence of abortion in cows and ewes, some deaths in ewes after abortion and a high
mortality rate due to enteritis in lambs younger than a few weeks of ages. In cattle, the first
signs may be fever and abortion, flowed several days later by diarrhea (Kahn, 2005; EFSA9,
2010). Pejcic-Karapetrovic et al. (2007) also found abortion in affected mice. These authors
found that pregnant mice became fatally susceptible to infections due to defective systemic
immunity.

Chronic enteritis: is a common form in pigs and adult cattle. There is persistent diarrhea,
severe emaciation and intermittent fever. The faeces are scant and may be normal or contain
mucus, casts or blood. Affected pigs are anorectic and lose weight (Kahn, 2005).
In Appendix I the potential adverse effects of Salmonella in animals are shown.
Humans
In humans, the high-risk populations for Salmonella infections include the young, old,
pregnant, transplant patients, and HIV-infected individuals. Pregnancy poses a high risk,
but it is unclear how maternal immunity to infection is altered (Pejcic-Karapetrovic et al. 2007).
Typhoid fever is a systemic disease caused by Salmonella typhi. The onset of this Salmonella
infection often gradual; the symptoms consist of fever, malaise, anorexia, headaches, and
muscle pain. Remittent fever is prominent. Either constipation or diarrhea may be present.
Respiratory symptoms, including cough or sore throat, may be prominent (Zettel et al., 1995).
The Merck Manual (Beers, 2006) also report central nervous system effects, slow heart
rhythm, exhaustion, enlargement of the spleen, decrease in the number of white blood cells,
anaemia, liver function abnormalities and proteinuria. Late in the disease intestinal lesions are
most prominent. Patient may suffer from severe intestinal bleeding and may die.
Non-typhoidal Salmonella infections, mainly caused by Samonella enteritidis, primarily
produce gastroenteritis, bacteremia (bacteria present in blood) and focal infection. Fever,
exhaustion and septicaemia are also symptoms described, however in a less severe form than
typhoid fever. Paratyphoid fever is a non-typhoidal Salmonella infection and is caused by three
serotypes of Salmonella enteritidis: Paratyphi A, Paratyphi B and Paratyphi C (Gupta et al.,
2008) is an emerging enteric fever (Gupta et al., 2008; Kanungo et al., 2008). Kanungo et al.
(2008) report treatment failure in India associated with increased mortality.
Zettel et al. (1995) have described, although rare, a patient with non-typhoidal Salmonella as a
cause of first-trimester pregnancy loss.
In Appendix I the potential adverse effects of Salmonella in humans are shown.
5. Severity of the potential adverse effects

The severity of contamination of food and / or feed with Salmonella is based on the worst case
scenario, as shown in table 7, and is based upon the potential adverse effects stated in
chapter 4 and Appendix I.
The severity of Salmonella toxicosis in animals is classified as high because:

In animals death can occur (Shivaprasad, 2000; Kahn, 2005);
Abortion has been reported in infected animals (Kahn, 2005; Pejcic-Karapetrovic et al.
2007; EFSA9, 2010).
The severity of Salmonella toxicosis in humans is classified as high because:

Mortality from typhoid and paratyphoid fever has been reported (Beers, 2006; Kanungo et
al., 2008);
Abortion caused by non-typhoidal Salmonella has been described (Zettel et al., 1995).
Table 7. Severity of Salmonella toxicosis

Severity
Low Medium High
Animals x
Humans x

6. Legislation and standards
This chapter focuses on legislation and standards applicable to feed.
There are numerous surveillance systems in the EU on Salmonella in feed of which some are
compulsory in certain MSs (EFSA1, 2009; Wierup and Hggblom, 2010). The GMP+ FSA
Scheme of GMP+ International has a strong focus on hygiene and Salmonella control and
monitoring, e.g. in the GMP+ B1, BA4 and BA13 standards (GMP+1, 2012; GMP+2, 2012;
GMP+3, 2012). In the GMP+ BA1 standard Salmonella product standards are stated (GMP+4,
2012).
In Norway a Salmonella surveillance system in feed has been established (Hofshagen et al.,
2010). The Belgian OVOCOM has stated an action level concerning Salmonella in feed
(OVOCOM1, 2011).
In the USA the Food and Drug Administration (FDA) has published the Notice of Availability
of Draft Compliance Policy Guide Section 690.800 Salmonella in Animal Feed in the Federal
Register in 2010. The draft CPG, when finalized, will help guide FDA staffs regulatory policy
relating to feed that are contaminated with Salmonella (US-FDA2, 2010). The American Feed
Industry Association has published Salmonella control guidelines for the American feed
industry (AFIA, 2010).
Besides the countries / continents mentioned above, several other Salmonella monitoring and
control programs in feed exist worldwide.
7. Methods of analysis
In GMP+ BA4 and BA13 requirements for sampling and analysis are stated (GMP+1, 2012;
GMP+2, 2012). In Appendix V these are categorised and summarised. Additionally other
recommendations, not being GMP+ requirements, are included.
For detailed information concerning method of Salmonella analysis, see Appendix VI.
Because of the sporadic nature of Salmonella contamination, most tests carried out for
monitoring purposes are negative. It is therefore valuable to monitor also indirect indicators
(EFSA6, 2008). Of these, Enterobacteriaceae counts are probably the most meaningful and
simplest to apply (Gradel et al., 2003; Jones and Richardson, 2004). The level of indicator
organisms is however not always linked with the risk of Salmonella contamination, e.g.
Salmonella originating from oilseed residue or pellet cooling systems may be present
regardless of the microbiological status of the feed in terms of faecal indicators or other
organisms. It is therefore not desirable to designate microbiological criteria for indicator
organisms but it is appropriate to refer to the voluntary use of microbial counts according to
requirements of feed manufacturers (EFSA6, 2008).
E. coli can be recommended as a reliable indicator organism for the potential presence of S.t.
in manure-fertilized soil (Natvig et al., 2002).
The ISO 21528:2004 standardised methods for the detection of Enterobacteriaceae include
the MPN (Most Probable Number) technique and colony counts. For direct enumeration the
pour plate method can be used.
Commercial Enterobacteriaceae identification systems are based on one of five different
technologies or a combination thereof. These include pH-based reactions that require from 15
to 24 h of incubation, enzyme-based reactions that require 2 to 4 h, utilization of carbon
sources, visual detection of bacterial growth, or detection of volatile or nonvolatile fatty acids
via gas chromatography (OHara, 2005).

8. Possible control measures
The HACCP system should be used to assess the risk of Salmonella in feed and consequently
the control measures needed to control this risk. All parts in the feed and food chain should be
included, starting from cultivation or husbandry.
Possible control measures are related to specific activities within each part of the feed and
food chain. However some control measures, e.g. concerning hygiene, are applicable in each
part of the feed and food chain.
In Appendix VII possible general Salmonella control measures are mentioned, applicable for
several part in the feed and food chain.
The general control measures can be complemented with the specific possible Salmonella
control measures for specific parts in the feed and food chain. These specific control
measures are included in the appendices:
Cultivation, harvesting, processing, transport and storage on farm, see Appendix VIII;
Husbandry, see Appendix IX;
Transport, see Appendix X;
Storage, see Appendix XI;
Processing of feed, see Appendix XII.
The Possible control measures should focus on products with a higher risk to contain
Salmonella are summarised in table 8.
Table 8. Products with a higher risk to contain Salmonella.

Manure is a possible source of Salmonella contamination of soil and water;
Any product fertilized with manure;
Concerning drinking water e.g. wells and water/tank vessels can be contaminated via vermin
Products in general
and their excrements of vermin or manure;

Any product in contact with contaminated irrigation water or water droplets;
Concerning soil, adherence of contaminated soil to vegetable surface is a risk. This can
occur during cultivation or in the process of harvesting (e.g. during reaping close to the soil);
Any product in contact with pests, other wild or domesticated animals or in contact with
their excrements;
Any product in contact with contaminated air and/or dust (e.g. fan-driven, dust,
fluff/down);
Any product in contact with Salmonella contaminated material (e.g. residues in production
lines, residues of previous cargoes etc.).
Any product not (heat/acid) treated in order to kill Salmonella
Products from the oil crushing industry:

Soy bean products or products containing soy beans products (e.g. soy bean meal
Specific products of
and expeller, soy protein concentrate, toasted soy beans);

vegetal origin
Rape seed products or products containing rape seed products (e.g. rape seed meal
and expeller);
Sunflower meal;
Palm kernel products or products containing palm kernel products (e.g. palm kernel
meal);
Other products of vegetal origin, e.g. cotton seeds, maize, maize gluten meal, rice products,
wheat bran, brewers grain, nuts and nut products and seeds (e.g. peanut products,
sesame seeds, pine nuts), herbs and spices. Or products containing these products.
Products of animal origin are of risk, specifically animal-derived proteins, e.g. meat and
products of
bone meal, poultry offal meal, fish meal and egg shells. Dairy by-products can also be of
Specific
animal
origin
risk, e.g. Category 3 milk, milk-based products and milk-derived products, raw milk and
raw milk-based products (incl. on-farm waste milk), raw colostrum and white water.
Products of animal origin destined for human consumption of risk are meat, poultry, eggs, milk
and fishery products.
Young animals have a greater susceptibility for Salmonella that may be due to the high
gastric pH, absence of a stable intestinal flora and limited immunity (Kahn, 2005).

9. References
1. AFIA (American Feed Industry Association), Salmonella control guidelines, 2010
2. Alali et al., Prevalence and distribution of Salmonella in organic and conventional
broiler poultry farms, Foodborne Pathogens and Diseases, Volume 7(11), 2010, pages
1363-1371
3. Alsop, An outbreak of salmonellosis in a swine finishing barn, Case report, Peer
reviewed, Journal of Swine Health and Production, Volume 13(5), 2005, pages 265-
268
4. Altamura et al., Splenectomy and sepsis: The role of the spleen in the immune-
mediated bacterial clearance, Immunopharmacology and Immunotoxicology, Volume
23(2), 2001, pages 153-161
5. Bailey and Cosby, Salmonella Prevalence in Free-Range and Certified Organic
Chickens, Journal of Food Protection, Volume 68(11), 2005, pages 2451-2453
6. Bailey et al., Sources and Movement of Salmonella through Integrated Poultry
Operations: A Multistate Epidemiological Investigation, Journal of Food Protection,
Volume 64(11), 2001, pages 1690-1697
7. Barak and Liang, Role of Soil, Crop Debris, and a Plant Pathogen in Salmonella
enterica Contamination of Tomato Plants, PLoS One, Volume 3(2), 2008, e1657
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APPENDIX I Potential adverse effects of Salmonella
3,4 2,3 2,3 2,3 3 3 3

Death Carcinogen Mutagen Reproductive Internal injury Neurological Immunological Organs
(physical
2,3
contamination)
Animals
x x x x x
Humans
x x x x x
3 3 3 3
Dermal and Respiratory Musculo- Cardiovascular Gastrointestinal Hematological Endocrine Bodyweight
3 3
ocular skeletal
Animals
x x
Humans
x x x x x
3
This potential adverse effect is classified as high severity for animals
4
This potential adverse effect is classified as high severity for humans

APPENDIX II Trends in most important Salmonella-serotypes in humans (RIVM1, 2011;
RIVM3, 2012).
2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011
Total number 2086 1591 2144 1626 1333 1668 1262 1648 1229 1446 1299
Serotype%total % % % % % % % % % % %
Enteritidis 43.2 44.4 55.2 47.2 35.9 37.5 36.8 34.3 33.0 35.3 29.3
Typhimurium 34.0 31.9 24.0 28.5 39.6 37.0 26.2 35.2 32.9 29.8 25.6
SI 1,4,5,12:i:- -- -- -- 0.1 1.8 4.9 5.4 5.8 7.0 9.4 20.2
(Para-)Typhi (A,B,C) ? 1.4 1.8 1.9 1.1 1.4 1.7 1.3 2.0 1.3 1.2
Infantis 1.2 2.0 1.3 1.4 1.4 1.1 1.0 0.8 2.7 1.2 1.0
Virchow 1.4 1.9 1.0 0.8 1.3 1.5 1.8 1.3 1.2 0.7 0.1
Brandenburg 1.3 2.1 0.9 1.2 1.8 0.5 0.6 0.4 0.3 0.4 0.3
Hadar 1.0 1.1 1.0 1.0 0.9 1.0 0.5 0.3 0.3 0.5 0.1
Bovismorbificans 1.3 0.9 0.5 0.6 0.2 0.3 0.2 0.5 0.2 0.3 0.5
Goldcoast 0.4 0.8 2.1 0.8 0.2 0.4 0.6 0.1 0.2 0.1 0.5
Newport 0.8 0.5 0.8 0.8 0.5 0.7 2.1 0.7 2.0 1.4 2.2
Panama 1.8 0.3 0.3 0.5 0.3 0.3 0.6 2.1 -- 0.5 0.3
Derby 0.5 0.7 0.8 0.6 1.0 0.4 0.6 0.2 0.7 0.6 0.8
Kentucky 0.2 0.7 0.3 1.3 0.8 0.7 1.5 1.2 1.1 1.0 1.1
Livingstone 0.6 0.0 0.5 0.4 0.3 0.1 0.3 0.1 0.2 0.1 0.1
Heidelberg 0.2 0.1 0.5 0.8 0.4 0.4 0.6 0.5 0.8 0.2 0.6
Dublin 0.6 0.3 0.2 0.4 0.2 0.5 0.5 0.4 0.2 0.3 0.6
Corvallis 0.1 0.1 0.1 0.4 1.3 0.5 0.6 0.8 0.8 1.0 1.0
Agona 0.2 0.5 0.2 0.5 0.1 0.3 0.7 0.2 0.3 0.3 0.4
Saintpaul 0.2 0.2 0.1 0.4 1.3 0.4 0.6 0.8 0.4 1.3 0.4
Schwarzengrund ? 0.1 0.1 0.1 0.1 -- 0.2 0.4 -- 0.3 0.2
Braenderup 0.2 0.5 0.3 0.1 0.4 0.4 0.2 0.3 0.1 0.5 0.5
Stanley 0.2 0.3 0.5 0.4 0.7 0.4 0.6 0.4 1.0 0.4 0.5
London 0.6 0.3 0.3 1.0 0.1 0.2 0.2 0.4 0.4 0.2 0.5
Manhattan ? 0.9 0.1 0.1 0.1 0.2 0.1 0.1 0.2 -- 0.1
Paratyphi B var. Java 0.4 0.4 0.1 0.2 0.1 0.2 0.8 0.6 0.7 0.6 1.2
Mbandaka 0.1 0.1 0.1 0.6 0.2 0.2 0.7 0.1 0.2 0.6 0.1
Oranienburg 0.7 0.3 0.2 0.2 -- 0.4 1.0 0.7 0.2 0.5 0.6
Montevideo 0.1 0.2 0.2 0.4 0.4 0.4 0.6 0.2 0.3 0.4 0.3
Senftenberg 0.1 -- 0.1 0.2 0.5 0.5 0.6 0.4 0.2 0.1 --
Blockley 0.2 0.3 0.2 0.1 0.5 0.5 0.1 0.2 0.2 0.1 --
Poona 0.4 0.1 0.1 0.3 0.2 -- 0.4 0.2 0.1 0.4 0.4
Anatum 0.2 0.2 0.2 0.4 0.5 0.1 0.5 0.4 -- 0.3 0.2
Muenchen 0.2 0.2 0.1 0.1 0.4 0.7 0.2 0.1 0.1 0.1 0.2
Muenster ? -- 0.4 0.1 -- 0.2 0.3 0.2 0.2 0.1 0.1
Weltevreden 0.2 0.1 0.0 0.1 0.1 0.1 0.8 0.3 0.3 0.4 0.3
Thompson 0.1 -- 0.1 0.0 0.2 0.2 0.2 0.4 0.2 0.5 --
Give 0.1 0.1 0.1 1.0 0.2 0.1 0.3 0.2 0.2 -- 0.2
Bredeney 0.3 0.3 -- 0.1 0.2 0.3 0.8 -- -- 0.1 ?
Napoli -- 0.1 -- 0.1 0.1 0.1 0.2 0.5 0.8 0.2 0.5
Mikawasima -- -- -- 0.2 -- 0.5 0.2 0.1 0.1 -- ?
SI 9,12:NM ? 0.4 0.2 0.2 0.1 -- 0.3 0.1 -- 0.5 0.1
SI 4,5,12:b:- 0.1 0.1 -- 0.1 0.2 0.2 0.1 0.4 0.7 0.8 0.3
SI 9,12:l,v:- 0.2 0.4 0.3 0.2 0.5 0.4 0.5 0.2 0.2 -- 0.3
Other serotypes 4.7 6.3 5.3 4.8 4.9 4.6 8.3 6.8 7.5 8.0 7.5
? = not reported

APPENDIX III Feed and food of vegetal origin of risk and Salmonella outbreaks in the
EU
Reported Salmonella outbreaks in the EU:

Reported (EFSA1, 2009) Salmonella outbreaks in the EU were in lettuce, tomatoes and
basil. In Germany and the Netherlands, Salmonella was detected in relatively high
proportions in ready to eat (RTE) sprouts (2.2% and 1.5% of the tested samples,
respectively). In 2011 a Salmonella outbreak in the Netherlands and Germany was related to
(bean) sprouts (RIVM3, 2012). In the EU RASFF Portal (DG SANCO, 2012) Salmonella
contaminated food and feed are registered. Several vegetal feed ingredients have been
reported contaminated with Salmonella, with varying serotypes. Most frequently reported
feed are soy bean meal and -expeller, soy protein concentrate, rape seed meal and
expeller, sunflower meal and maize gluten meal. Herbs and spices were also frequently
reported to be Salmonella contaminated. Other feed reported are sunflower expeller, palm
kernel expeller and linseed expeller, yeast. Food frequently reported are nuts and nut
products and seeds (e.g. peanut products, sesame seed and pine nuts).
Salmonella critical feed and food of vegetal origin:

Despite the sampling uncertainties, ingredients used for feed have been shown in several
studies to commonly be contaminated with Salmonella. The EFSA6 (2008) focuses on
vegetal protein and distinguishes non-processed and processed products. Data provided by
Defona, Norwegian feed supplier, cited by EFSA6 (2008) show that 30% of the dust samples
non-processed soy beans, tested positive for Salmonella. The soybeans were imported
mainly from South America to Norway during 1994-2007.
Concerning processed vegetal protein, several examples also illustrate that products from
the crushing industry are often contaminated with Salmonella. In the recent past soy bean
meal and expeller and toasted soy beans have been classified as Salmonella critical within
the GMP+ system of GMP+ International. As an example, in a Dutch report feed from 2002
and 2003, 3.2% and 6.7% respectively of Brazilian soy bean meal were positive for
Salmonella (PDV2, 2004). In the years 2008, 2009 and 2010 reported Salmonella positive
samples in Brazilian soy bean meal samples were 1.0%, 0.3% and 0.5% respectively (data
originating from the DOS-database of GMP+ International), indicating a lower incidences as
shown in table 3. In Sweden 14.6% of imported consignments of soy bean meal were found
to be contaminated by Salmonella during 2004-2005 and when considering only imports
mostly from South America the level was approximately doubled (Wierup, 2006).
Available data from the EFSA zoonoses report 2005 also support oil seeds e.g. soybean and
rapeseed products, as a risk factor for introducing Salmonella into the feed chain (EFSA7,
2006). In the recent past rape seed meal and expeller have been classified as Salmonella
critical within the GMP+ system of GMP+ International. (EFSA6, 2008). In Sweden rape seed
products are classified as high risk (SVA, 2011). The Product Board Animal Feed (PDV5,
2007) reported a contamination rate for rape seed meal and expeller of 6.8 and 3.4% for the
years 2005 and 2006 respectively. In the years 2008, 2009 and 2010 reported Salmonella
positive samples in rape seed meal samples were 1.5%, 0.7% and 1.1% respectively (data
originating from the DOS-database of GMP+ International), indicating a lower incidences as
shown in table 3. Wierup and Hggblom (2010) report that 10% of the in Sweden imported
rape seed meal in 2004 and 2005 was found to be contaminated with Salmonella.
In the recent past sunflower meal has been classified as Salmonella critical within the
GMP+ system of GMP+ International (PDV1, 2003; PDV2, 2004; PDV3, 2005; PDV4, 2006). In
the EU RASFF Portal (DG SANCO, 2012) Salmonella contaminated sunflower meal has
been reported.

Salmonella is also reported from palm kernel products (PDV2, 2004; PDV4, 2006; PDV5,
2007). In Norway palm kernel meal was in 2004 and 2005 classified as high risk feed
(Wierup and Hggblom, 2010).
In a study of Torres et al. (2011) it was found that of the feed used, cotton seeds were
identified as having the highest odds of Salmonella contamination
Grain is seldom found to be contaminated unless as a result of contamination during storage

and transport. However, studies show that grain can also be contaminated, especially if it
originates from areas where Salmonella is common in wildlife or local livestock (EFSA6,
2008). In Norway maize was in 2004 and 2005 classified as high risk feed (Wierup and
Hggblom, 2010). In Sweden rice products are classified as high risk (SVA, 2011).
In the recent past wheat bran has been classified as Salmonella critical within the GMP+
system of GMP+ International. In the EU RASFF Portal (DG SANCO, 2012) maize gluten
meal was also reported to be contaminated with Salmonella. Green et al. (2010) studied risk
factors associated with detection of Salmonella from cattle in U.S. feedlots. Concerning feed
the authors found that the inclusion of maize gluten and brewers grain in pen rations were
associated with an increase in likelihood of Salmonella detection.
GMP+ International states that at the moment of publication of this fact sheet, no feed is
classified as Salmonella critical (GMP+1, 2012). This is the result of control measures taken
by several GMP+ participants in the feed/food chain. However this situation might not reflect
the situation in every part in the world. In some cases a specific origin of the feed was
classified as of risk. UK data also records an improving trend in the contamination of oilseed
meals and products from 4.3% in 2002 (VLA1, 2003) to 0.9% in 2009 (VLA2, 2010).
The industry based data from 2001 and 2010, as shown in table III.1 (PDV1, 2003; PDV2,
2004; PDV3, 2005; PDV4, 2006; PDV5, 2007; PDV6, 2008; for 2008-2010 data in the DOS-
database of GMP+ International were used) reports incidences of Salmonella contaminated
samples in feed of vegetal origin (not being liquid by-products). For more information
concerning these data is referred to the references cited. Mind that in some cases the
number of samples analysed is small and the reported Salmonella incidence might not reflect
the actual situation.
Table III.1. Industry Salmonella data feed of vegetal origin (not being liquid by-products) GMP+ certified (GMP+
1 2 3 4 5 6
FSA scheme) companies (PDV , 2003; PDV , 2004; PDV , 2005; PDV , 2006; PDV , 2007; PDV , 2008; for
2008-2010 data in the DOS-database of GMP+ International were used).
Feed 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011
% % % % % % % % % %
Bakery by-products 0.3 0 0 0 0 0 0.6 0.9 0 0
Cereals and cereal

by-products:
Barley and barley 0 0.6 0 0 1.6 0 0 0 0 0.3
by-products
Maize 0 0.6 0.7 0.2 0 0.1 0.3 0 0 0
Maize by- 1.2 0.6 0.3 1.6 0 0 0.7 0.7 13.3 10.0
products
Oat and oat by- 13.6 0 0 0 0 0 0 0 0 0
products
Rice and rice 0 1.9 3.7 0 0 0 0 0 0 4.3
products
Rye and rye by- 0 0 0 0 0 0 0 0 0 0
products
Triticale and 0 0 0 1.5 0 0 0 0 0 0
triticale by-
products
Wheat and wheat 1.1 0.7 0.2 0.2 0 0.3 0.1 0 0 0.1

Feed 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011
% % % % % % % % % %
by-products
Citrus pulp 0 0 0 0 0 0 0 0 0 0
Linseed expeller 3.7 2.2 7.5 8.9 - - - - - -

Linseed products - - - - 0 0 0 2.2 0 0
Palm kernel 0 0 0 0.8 0 0 0 0 0 0

expeller
Palm kernel + - - - - 7.1 0 0 0 - 0
extracted
Palm kernel 0 10.5 0 0 - - - - - -
extracted
Potato products 2.6 0 0.9 0 0 0 0 0 0 0
Pulses 0.8 0 0 0 0 0 0 0 0 0
Rape seed 0 - 0 8.7 0 - 0 0 0 0

Rape seed, 12.0 7.0 1.5 0.7 1.1 1.6
extracted
11.8 6.8 3.4 1.1
Rape seed, - 1.7 0.5 0.2 0.3 0.4
expeller
Roughage (grass / 8.5 0 0 2.2 0 0 0 0 0 0

alfalfa)
Soy bean, raw - 3.1 0 6.0 0 0.7 0 - 0 0

Soy bean, expeller - - 6.3 3.6 3.2 0 0 0 4.6
Soy bean, 2.0 4.6 5.2 3.4 1.3 2.6 1.8 1.1 0.9 0.6
extracted
Soy bean, other 1.7 0.7 2.6 1.0 2.4 0.2 0.4 0.6 0 0.6
1
products
Soy bean, heated 2.0 3.3 4.9 4.3 2.0 1.3 0.4 1.9 1.3 1.9
Sunflower seed, 4.4 3.2 4.8 3.5 1.4 0.9 0.8 0.2 0.4 0.5
extracted
Sunflower seed + - - - - 15.4 0 0 0 11.8 0
expeller
Sunflower seed, 0 0 3.8 0 - - - - - -
expeller
1
not being soy oil or soy oil fatty acids
Based on the data presented in table III.1 it can be concluded that soy-, rape, maize and
sunflower based products had the highest Salmonella incidences in the period of 2001-2011.

APPENDIX IV Feed and food of animal origin of risk
Certain feed of animal origin is of risk concerning Salmonella, specifically animal-derived

proteins. When allowed as ingredients in feed, mammalian meat and bone meal (MBM)
and poultry offal meal were frequently contaminated by Salmonella, a logical consequence
of the risk from the rendering of animals infected with Salmonella. The risk for the Salmonella
contamination attributes to in-house contamination in the rendering plants and
recontamination following the heat treatment process (EFSA6, 2008). Monitoring by the EU
feed industry also confirms that animal protein products frequently are contaminated by
Salmonella; during 2005 14.9% of 94 samples were contaminated, 8.3% of 72 samples
during 2006 (PDV5. 2007) and in 2011 2.6% of the 38 samples were contaminated with
Salmonella (data originating from the DOS-database of GMP+ International). The EFSA
(2012) reports a Salmonella contamination level of 0.6% in meat and bone meal in 2010.
Fish and seafood are also susceptible for Salmonella contamination (Heinitz et al., 2000). A
major Salmonella Thompson outbreak occurred in the Netherlands in 2012. The cause was
traced back to smoked salmon of one producer. Insufficient cleaning and disinfection of
equipment may have resulted in an increase and persistence of the contamination of the
production line (Friesema et al., 2012), especially since S. Thompson has been reported to
easily form a persistent biofilm (Warner et al., 2008; Patel and Sharma, 2010; Sha et al.,
2012). According to the producer a design of the packaging material is responsible for the
outbreak. Salmonella Thompson does not specifically seem to host on fish products or other
products of animal origin, however the serotype has been found on products of animal origin;
beef (Shapiro et al., 1999), beef and poultry carcasses (Madden et al., 2001; Sakaridis et al.,
2011) and pet treats (US-CDC, 2006). A link with animal feed was not established or was not
studied in these publications. The serotype was not reported in the top 25 serotypes found in
animal feed in the U.S. between 2002 and 2009 (Li et al., 2012). Concerning fish meal or
other animal feeds of animal origin no S. Thompson data were found.
Fish meal has the potential for the spread of Salmonella although fish meal seems to be
somewhat less contaminated than other animal derived protein feed according to the EFSA
zoonoses report from 2005 (EFSA7, 2006). As for MBM the risk of Salmonella contamination
is found to be attributed in-house infection in the rendering plants and recontamination
following the heat treatment process (EFSA6, 2008). The EFSA reports data on Salmonella
in fish meal in EU Member States (MSs) derived from different surveillance programmes.
The observed levels of contamination in fish meal ranged from 1.9% to 2.9% in 2006 and
2007 respectively (EFSA9, 2010). In 2009 0.7% of the fish meal tested, was found to be
contaminated with Salmonella (EFSA10, 2011). However in 2010 a marked increase was
observed in reports of Salmonella contaminated fish meal, being 9.1 % (EFSA12, 2012)
Industry based data from 2001 and 2010, as shown in table IV.1 (PDV1, 2003; PDV2, 2004;
PDV3, 2005; PDV4, 2006; PDV5, 2007; PDV6, 2008; for 2008-2011 data in the DOS-database
of GMP+ International were used) reports a range of 0% to 3.5%, as shown in table 4.
There is a potential risk for the spread of Salmonella by feeding animals also by some dairy
by-products. In particular ready-to-use Category 3 milk, milk-based products and milk-
derived products, and raw milk and raw milk-based products, unless they have been
subjected to further treatment, as prescribed in EU legislation (EFSA8, 2006). Several
authors found raw milk to be contaminated with Salmonella (Van Kessel et al., 2004; Jayarao
et al., 2006). Houser et al. (2008) conducted a survey of the occurrence of Salmonella in raw
colostrum and found that 15% of the samples were contaminated with Salmonella. Raw milk
and raw milk based products, including colostrum, are not approved in the feed industry.
However feeding of these products obtained, kept, disposed of or used on the farm of origin
is known. The 2002 US NAHMS (National Animal Health Monitoring System) Dairy Survey
indicated that more than 85% of dairy farms in the United States feed unpasteurized waste
milk to their neonatal calves (USDA, 2002). Although cost-effective, this practice can lead to
increased calf morbidity and mortality due to ingestion of pathogenic agents, including

Salmonella (Stabel et al., 2004). Related to diary products, the EFSA8 (2006) mentions
white water5 as Salmonella risk ingredient.
Egg shells which are not heat treated are known to be frequently contaminated with
Salmonella.
The industry based data from 2001 and 2011, as shown in table IV.1 (PDV1, 2003; PDV2,
2004; PDV3, 2005; PDV4, 2006; PDV5, 2007; PDV6, 2008; for 2008-2011 data in the DOS-
database of GMP+ International were used) reports incidences of Salmonella contaminated
samples in feed of animal origin. For more information concerning these data is referred to
the references cited. Mind that in some cases the number of samples analysed is low and
the reported Salmonella incidence might not reflect the actual situation. The animal protein
products are products not to be used in feed destined for farmed animals (food producing
animals). Animal protein products, including offal meal, and fish products (e.g. fish meal) had
the highest Salmonella incidences.
Table IV.1. Industry Salmonella data feed of animal origin GMP+ certified (GMP+ FSA scheme) companies
1 2 3 4 5 6
(PDV , 2003; PDV , 2004; PDV , 2005; PDV , 2006; PDV , 2007; PDV , 2008; for 2008-2011 data in the DOS-
database of GMP+ International were used).
Feed 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011
% % % % % % % % % %
Animal fat (warm-blooded 0 0 0 0 0 0 0 0 - 0

+ fish))
Animal protein 5.3 5.3 3.4 14.9 8.3 9.4 12.5 6.3 13.0 2.6
products (warm-blooded)
Dairy products 0 0 0 0 0 0 0 0 0 1.1

Milk powder, 0 0 0 0 - - - - 0 0
products
Whey and whey 1.1 0 0 0 0.2 0 0 0 0 0
products
Fish meal 3.5 1.2 0.8 0.8 0.8 0.4 - - - -

Fish products (incl. fish - - - - - - 0 0.4 3.1 0
meal)
Egg products 0 0 0 0 0 0 0 0 0 0
Egg shells 27.3 - - - 0 0 0 0 0 0
(heated) (heated) (heated)
Based on the data presented in table IV.1 it can be concluded that animal protein products
(e.g. meat and bone meal) and fish products (e.g. fish meal) had the highest Salmonella
incidences in the period of 2001-2011.
5
White waters come from washing operations at dairy facilities and are separated from those coming from the
cleaning and disinfection of the equipment. They may also come from washing operations of dairy facilities before
heat treatment, including from trucks delivering milk to dairy plants, and will contain raw milk. In some cheese
industries white waters and whey may be mixed before rejection. White water is, in some cases, heat-treated and
then dehydrated in spray dryers before being sent to a farm for animal feeding, or it may be transported as liquid
feed

APPENDIX V Sampling
In GMP+ BA4 and BA13 requirements for sampling are stated (GMP+1, 2012; GMP+2, 2012).
Additionally other recommendations, not being GMP+ requirements, are stated below in this
appendix:
Sampling
Confidence in the results of Salmonella testing will depend on the number of samples units
tested, whether or not there is a homogeneous distribution of the target organism in the lot
and whether the sampling is performed randomly (EFSA6, 2008).
Sampling material
Samples can be taken using disinfected (with 95% alcohol or another bactericidal agent)
or sterile scoop, hand scoop or sample drill (GMP+2, 2012);
Use sterile sampling containers (e.g. bags, pots, bottles);
Avoid the us of thin bags (AFIA, 2010);
Avoid clear or see-through containers since they can trap solar energy (AFIA, 2010) and
produce a possibly false-negative result of analysis;
Use sterile gloves, disinfect hands (GMP+2, 2012);
Sampling location
Every attempt should be made to collect samples that reflect or represent the entire load
or batch of feed. When sampling a previously loaded vehicle or material from storage,
samples can be taken from each of 10 different locations. For bulk shipments, take a
sample at varying intervals of time as material is unloaded or loaded so the sample
represents the contents of the load (AFIA, 2010);
In GMP+ Appendix BA4 the sampling locations and frequencies for feed for which
Salmonella monitoring is obliged for GMP+ certified participants, can be found (GMP+1,
2012);
Delivered feed should preferably be sampled during loading or unloading of the transport
entity. If this is not possible then from the stationary vehicle auto where the whole load
must be accessible. If the product is bagged then a sample can be taken during bagging
(GMP+2, 2012).
Sampling technique
Do not cough, sneeze or talk during sampling (GMP+2, 2012);
Samples must be taken such that contamination, for example by rain or dust, of the
samples or containers in which the samples are stored is prevented (GMP+2, 2012);
Take measure to avoid infection from clothing, hair etc. during sampling (GMP+ 2, 2012);
Keep sampling containers. Open as shortly as possible and with the opening turned
upwards at an angle of 45 (GMP+2, 2012);
Do not touch the insides of sampling containers, incl. covers, and the sampling tools with
the hands (GMP+2, 2012);
Hold scoops etc. by the handles (GMP+2, 2012);
Avoid sampling by pouring out. If this cannot be avoided then disinfect the edge over
which pouring will be done prior to used (GMP+2, 2012);
Avoid contact with heat, sunlight, damp and equipment (GMP+2, 2012);
The sample size amounts to at least 60 grams, which is sufficient for a duplicate
determination (GMP+2, 2012).
Sample sealing, storage and consignment
The sample should be labeled such that it can easily be identified. At least the following
records must be made, if applicable, on the sample using a clearly linked form of
registration: date of sampling, product identification, batch identification, sampler,
supplier, production unit where the sample was taken (GMP+2, 2012);
The sample must be kept in such a way that damage to and deterioration of the sample
is avoided (GMP+2, 2012);
The sealing should be such that opening the sample is inevitably leads to an irrevocable

Sampling
break in the seal on the sample (GMP+2, 2012);

Deliver samples of liquid by-products to laboratory within 24 hours. Other samples must
be sent within two working days (GMP+2, 2012).
Sampling frequency
In GMP+ Appendix BA4 the sampling frequency for feed for which Salmonella monitoring
is obliged for GMP+ certified participants, can be found (GMP+1, 2012).

APPENDIX VI Methods of Salmonella analysis
Methods of Salmonella analysis
In addition the sensitivity and specificity of the used testing method has to be taken into
account. Growth-based isolation and identification methods using enrichment and selective
media are used as the primary means to detect Salmonella in the feed chain. The low water
activity of most feed creates an environment where the bacterial cells are strongly
dehydrated and thus the isolation method must be able to give injured and stressed cells the
possibility to recover and multiply (EFSA6, 2008).
In GMP+ Appendix BA 13 a sampling protocol is stated for microbiological examination
(GMP+2, 2012).
In general there are four methods for Salmonella detection, the cultural, immunological,
molecular and quantitative method.
Cultural method
The international standard cultural method for detection of Salmonella is ISO 6579 (EFSA6,
2008). Another culture-based method used for Salmonella in feed is the NMKL-71 method
(Koyuncu et al., 2010). A major drawback of the culture methods is the time requirement
because most protocols use 5-7 days. Although direct plating may give rapid preliminary
results the low levels of Salmonella present in feed require selective enrichment of the
samples (EFSA6, 2008). Koyuncu and Hggblom (2009) compared three cultural methods for
the detection of Salmonella in feed, being the NMKL71-method, the Modified Semisolid
Rappaport Vassiliadis method (MSRV) and the international standard method (EN ISO
6579:2002). In conclusion, the cultural methods were shown to be surprisingly equivalent in
terms of accuracy, sensitivity and specificity for different kinds of feed and serotypes,
respectively. An interesting observation was the differences in detection levels between
different feed.
Immunological methods
Immunological methods generally apply enzyme-linked immune absorbent assay (ELISA). In
ELISA assays, enzyme linked mono- or polyclonal antibodies are used to detect antigens
from Salmonella. A main advantage is that those methods are more rapid than most cultural
methods and also possible to automate. In combination with magnetic beads these
techniques can also enhance the isolation of Salmonella from large samples. A drawback is
that cross reactivity with antigens in related bacteria may cause false positives. The
immunological methods may also be less effective for detecting stressed or damaged
bacteria (EFSA6, 2008).
Molecular methods
Molecular methods for the detection of Salmonella include conventional PCR and Real-time
PCR. The PCR technique is based on the detection of specific DNA sequences in genetic
material (EFSA6, 2008). Several commercial kits are available, that are capable of detecting
Salmonella with high specificity (Maciorowski et al., 2005). A major obstacle is that the small
numbers of salmonellae in feed present. The concentration of salmonellae in feed is
generally considered to be below the established detection level of the PCR assay (Lfstrm
et al., 2004). Another problem is that many feed contains substances that may be inhibitory
of the PCR reaction (Lfstrm et al., 2004). Koyuncu et al. (2010) compared commercial
PCR-based Salmonella enterica detection methods with culture-based methods and found
that many PCR positives could not be confirmed by Salmonella isolation and for that reason
the evaluated methods were found to be suitable only when rapid results are paramount.
Nevertheless, PCR-based methods cannot presently replace culture-based method when
typing information is required.
Quantitative methods
Quantitative methods are developed. The present methods for isolation of Salmonella give
qualitative results (absence/presence). Using serial dilutions it is possible to estimate the
most probable number (MPN) of bacterial cells in a sample. The traditional MPN method is

Methods of Salmonella analysis
labour-intensive and not suitable for large scale, however, simplified methods based on
microtiter plates are being developed (EFSA6, 2008).

APPENDIX VII Possible general control measures
In GMP+ B1 standard general control measures and requirements are stated which should
be complied (GMP+3, 2012). Additional recommendations for general control measures are
stated below in this appendix.
Personnel and other people
Each employee should be required to report to work in clean clothes and maintain
adequate, daily personal hygiene. It is highly recommended that employers provide clean
uniforms for use by the employees. The uniforms, footwear and other garments used in
the facility should stay on site and not be allowed to be taken home by the employees. If
reusable garments are used, the facility should provide for proper sanitizing (AFIA, 2010);
Persons suffering from communicable illness should not be allowed to work in the facility
(AFIA, 2010);
Every person must wash their hands with soap and water after each use of the restroom
and break facilities (AFIA, 2010);
Procedures should be in place to ensure that all visitors to the site, including staff,
contractors, transport operators and customers, are aware of the potential impact of their
actions on all aspects of product safety (AFIA, 2010);
Avoid unnecessary foot traffic from outside sources in all facility areas because
Salmonella contamination may be carried on the feet and shoes. Pay particular attention
to foot traffic from such areas as barns, stock pens and stock trucks to all facility areas
(AFIA, 2010);
Provide for sanitizing of footgear if deemed necessary and practical (AFIA, 2010);
Facilities
Facilities for employees can include the following (AFIA, 2010):

o Separate facilities for workers on the pre-processing side and workers on the post-
processing side of the plant;
o Adequate facilities for showering and dressing so as to minimize contamination carried
on clothing, shoes or the person;
o Adequate lavatory and toilet accommodations.
Sanitation and cleaning
Dust, dirt, organic material (AFIA, 2010) and residues of feed or food can be a major medium
for the growth of Salmonella, which can contaminate feed or food. The accumulation of dust,
dirt and feed or food residues must therefore be avoided as much as possible.
The following applies to all areas:
Cleaning and sanitizing procedures should be appropriate for each specific piece of
equipment or area (AFIA, 2010);
Cleaning should include the following: the interior and exterior of production machinery
as well as ceilings, roof structure, wall cavities, ledges or rafters (AFIA, 2010). Dust
extractors / collectors and coolers should be cleaned regularly (FEDIOL, 2009);
Magnets should be cleaned at regular intervals to prevent feed from building up at these
points (AFIA, 2010);
Dry cleaning of spillages by sweeping and/or vacuuming is preferable to wet, as water
contributes to Salmonella growth. If the use of water is necessary, appropriate water
temperatures and sanitizing steps should be specified (and appropriate to the area being
cleaned) (AFIA, 2010).
Settled dust should be removed using a vacuum cleaner rather than sweeping. Vacuum
cleaners should be dedicated to either pre- or post-process and should not be used in
other locations. Vacuum collection of dust is preferable to using compressed air, which
would increase air-borne dust. Employees should avoid using compressed air whenever

Sanitation and cleaning
possible to remove dust from equipment or from clothing of personnel (AFIA, 2010);
Decontamination after Salmonella detection
Once a Salmonella contamination has been detected, decontamination should take place. If
decontamination measures are inadequate, Salmonella organisms may adapt to the stress
conditions and become more resistant to control efforts (AFIA, 2010).
A procedure should be in place in case of the detection of Salmonella.
In some instances, equipment may need to be partially disassembled to allow access for
cleaning (AFIA, 2010);
When partial disassembly is not a solution, dry flushing may be an option. The flush
should be isolated, discarded and not re-used within the facility (AFIA, 2010);
In case of a Salmonella contamination in feed the cause of the contamination should be
determined. Appropriate measures should be taken to eradicate the cause. A procedure
should be in place describing how to handle Salmonella contaminated products.
Pest control
An effective pest control program should be in place. Pests include rodents, birds,
insects, but also wild and domesticated animals (e.g. dogs and cats);
Keep the grounds surrounding the facility well drained and free of unnecessary
vegetation, such as weeds and high grass (AFIA, 2010);
Keep all areas within and around the plant free from accessible waste and trash;
Minimize dirt, dust, spilt feed and other organic materials and clean up spills promptly
(AFIA, 2010);
Unused equipment should be stored in a manner that eliminates pest infestation (AFIA,
2010);
Dust control and air flow
Dust is inherent in feed manufacturing. However, dust is a primary vehicle by which

Salmonella can be transmitted within the facility. Therefore, measures taken to control and
minimize dust are crucial to the success of any Salmonella control program.
Ventilation within the production area should provide sufficient air exchange to prevent
accumulation of steam, condensation or dust and to remove contaminated air (AFIA,
2010);
Air drawn into the production area should not be taken from areas likely to be
contaminated (AFIA, 2010);
The control of air flow (and the dust contained in that air) should be designed so that
contamination does not spread from raw material areas into finished product areas of the
plant (AFIA, 2010);
Equipment has appropriate dust extraction capability (AFIA, 2010);
All processing and handling equipment should be constructed and installed in a manner
to minimize leakage, spillage and dust accumulation (AFIA, 2010);
Dust collection systems should be adequate to control dust and to aid in keeping the
plant clean (AFIA, 2010);
Dust extractors / collectors should be cleaned regularly and condensation should be
prevented where possible (FEDIOL, 2009);
Remove settled dust with a vacuum cleaner equipped with a highly efficient filter rather
than sweeping. Avoid using compressed air to remove dust from equipment or from
clothing or shoes of persons (AFIA, 2010);

Dust control and air flow
Dust from cleaning, maintenance and overhaul of equipment such as extractors,

cyclones and filters, should not re-enter the finished meal stream (FEDIOL, 2009), they
should be disposed of (AFIA, 2010);
The air used for cooling pelleted or extruded products is of particular concern, since
these products will normally be Salmonella-negative as a result of the temperatures and
pressures involved in these processes. While use of internal air is not preferable in
cooling, but commonly used in pelleting facilities, air filtration and dust control should be
in place to reduce possible recontamination of pelleted or extruded feed (AFIA, 2010).
Moisture control
The low water activity level found in most dry ingredients and finished products usually result
in severely dehydrated Salmonella bacteria. It is only when there is adequate moisture,
temperature and growing conditions that these stressed Salmonella bacteria recover and
multiply. Moisture is critical for Salmonella growth (AFIA, 2010).
The areas should be designed for wet weather operation, so that the loading and
unloading of feed occurs without significant water damage to the feed (AFIA, 2010);
Roofs, ceilings and walls should be leak-proof. Construct storage-area walls and floors in
such a manner as to keep out moisture (AFIA, 2010);
Keep feed dry at all times. (AFIA, 2010);
Avoid or correct conditions conducive to the formation of condensation in buildings,
equipment (AFIA, 2010) and storage and transport entities;
Pellet coolers should be operated in a manner to prevent condensation on interior
surfaces that encourage Salmonella growth and should be regularly monitored for
contamination (AFIA, 2010);
Any wet material should be disposed of as waste or recycled through effective heat-
processing step(s) (AFIA, 2010).

APPENDIX VIII Salmonella control measures: cultivation, harvesting, processing,
transport and storage of feed on farm.
Specific requirements and control measures related to feed ingredient cultivation are stated
in the GMP+ B6 standard (GMP+5, 2012). This standard should be complied with. Additional
recommendations for general control measures are stated below in this appendix.
Cultivation, harvesting, processing, transport and storage on farm
Use the HACCP system to assess the risk of Salmonella in feed. Besides others, the
following items should be addressed:
At the level of cultivation, contamination with Salmonella is possible through the
spreading of contaminated fertilizers (slurry, manure, waste sludge etc.) on the pasture /
fields, ingredients and by-products (EFSA6, 2008). Possible control measures are:
o Storage of the fertilizer for more than 2 months, without any new influx (EFSA6,
2008);
o Composting (EFSA6, 2008);
o Ploughing in after spreading fertiliser (EFSA6, 2008);
o Increasing the time allowed between spreading of the fertilizer and the animal grazing
or crop harvesting (EFSA6, 2008);
o Heat treatment of the fertilizers before used (EFSA6, 2008);
o Treating fertilisers with the addition of lime (EFSA6, 2008).
Water used for irrigation of crops should not be contaminated with manure / excrements
of pests, other wild or domesticated animals;
When harvesting avoid taking up soil contaminated with Salmonella with the crop;
Drying of crop, if harvested crop is too moist. Dry up to a moist content unfavourable for
the growth of Salmonella. Dried crop should be cooled up to a temperature as low to
prevent condensation and heating during storage;
Transport: avoid using transport entities also used for non-feed purposes (e.g. transport
of manure and animals);
Transport: regularly clean transport entities. If wet cleaning is needed, any residual water
after wet cleaning should be removed and all contact surfaces should be dry (AFIA,
2010);
Storage: maintain good hygiene during storage (EFSA6, 2008);
Storage: control the moisture of feed during its storage (EFSA6, 2008);
Storage: proof the basement, cover the silo in order to prevent contamination by pests,
wild and domesticated animals (EFSA6, 2008);
Storage: regularly monitor (visually) storage entities if the control of pests, wild and
domesticated animals is effective (EFSA6, 2008);
Storage: regularly clean storage entities (EFSA6, 2008);
In the event of Salmonella contamination corrective measures will be taken immediately.
Salmonella monitoring programs:

Implement various Salmonella monitoring programs, as stated hereafter. For more

information concerning sampling and Salmonella analysis, see chapter 7 of this fact sheet. In
GMP+ Appendix BA13 a sampling protocol is stated for microbiological examination (GMP+2,
2012). Several types of monitoring programs are suggested:
Implement a Salmonella monitoring program, if applicable, meeting the requirements
stated in applicable cultivation quality systems (e.g. Eurep-GAP). Concerning Salmonella
monitoring programs for the transport and storage of feed is referred to appendices X
and XI respectively. For specific information concerning Salmonella monitoring in
cultivation, see below;
Implement a Salmonella monitoring program on the following products, if applicable,

Salmonella monitoring programs:
which were determined of risk by the HACCP study: e.g. sowing seeds, irrigation water,
soil, fertilizer, manure, compost materials, residues and spills in specific areas;
Implement a Salmonella monitoring at critical points in cultivation, as is determined using
the HACCP system. Also include machinery and equipment in the monitoring program. A
representative number of samples should be taken and examined from the critical points.
Critical points where samples can be taken are e.g.:
o Dust from ledges inside and the top of feed silos / bins;
o Machinery and equipment in direct contact with feed;
In the event of Salmonella contamination corrective measures will be taken immediately,
see section Decontamination after Salmonella detection in Appendix VII.

APPENDIX IX Salmonella control measures: husbandry.
Husbandry
Use the HACCP system to assess the risk of Salmonella in feed during husbandry. Besides
others, the following items should be addressed:
Food producing animals are kept and certified according to the standards stated in a
quality program (e.g. IKB, KKM, QM-Milch etc.) addressing and controlling hygiene and
Salmonella;
Limit the number of visitors, respect the hygiene rules (farmer, visitor) and utilise special
clothes (EFSA6, 2008);
Home mixing of feed: the home mixing of feed should be certified in compliance with a
quality program addressing hygiene and Salmonella;
Home mixing of feed: for convenience the milling facilities are normally close to livestock
buildings and common vehicles such as tractors may be used in the mill and around the
farm, both delivering the feed to various parts of the farm and for other tasks including
harvest and livestock manure handling (EFSA6, 2008). Cleaning procedures of vehicles
and equipment used for milling activities should be in place;
In case of buying feed at third parties, suppliers can be selected based on their
implementation of programs for the control of Salmonella (EFSA6, 2008) or control of
hygiene;
Pests and other wild and domesticated animals: there is a significant chance of cross-
contamination of storage facilities (e.g. flat stores) for cereals and other feed by wild
birds, rodents, insects, feral cats and other animals whose movement is not controlled.
Ideally all feed used on farm should be stored in sealed bulk bins or in bags held in
rodent and bird-proof enclosures. Unfortunately this is rarely the case and in conclusion it
is important that feed is stored in a way that prevents and controls introduction,
multiplication or persistence of Salmonella. Proof the basement and cover the silo
(EFSA6, 2008).
Feed fed should be dry (if applicable), without organoleptic abnormalities (e.g. normal
smell and temperature, absence of mould);
Feed: no feeding of raw milk or by-products of raw milk (e.g. colostrum, waste milk, whey
of cheese production on-farm);
Water: food-producing animals should have access to clean drinking water (e.g. mind
when well water possibly contaminated with manure / excrements of pests and wild and
domesticated animals);
Water: use of authorized acids in drinking water (Van Immerseel2 et al., 2006);
Feeding system: avoid stepping on feed and feeding feed on walkways (EFSA6, 2008);
Feeding system: avoid the possibility of manure, litter, water, soil, slurry or sewage to get
in contact with feed (EFSA6, 2008);
Feeding system: use clean and dry buckets and teats for bucket feeding (EFSA6, 2008).
Group feeding may contribute to the spread of microorganisms (Hepola, 2003);
Feeding system: liquid feeding, particularly when whey is fed, or where fermentation is
controlled using a starter culture and controlled temperature conditions, may reduce the
risk of intestinal carriage of Salmonella (EFSA6, 2008);
Feeding system: leave no residues in the feeding system after feeding;
Feeding systems: open systems attract birds, rodents, insects and pets. Feed and feed
troughs might get contaminated via direct contact or via excrements (EFSA6, 2008). A
pest control program should be in place: see section Pest control. For possible control
measures concerning storage of feed in general: see section Storage and transport;
Feeding systems: cleaning procedures should be in place for feeding systems, including
equipment and machinery used.
Pig feeding systems with their complex of troughs, hoppers and pipes have been shown
to be particularly difficult to clean and disinfect effectively because of inaccessible

Husbandry
surfaces and pooling of wash water. This contamination may be responsible for carry-
over of Salmonella between batches of pigs (EFSA6, 2008);
If wet cleaning is needed, any residual water after wet cleaning should be removed and
all contact surfaces should be dry (AFIA, 2010);
Salmonella monitoring programs: husbandry

2012). Several types of monitoring programs are suggested:
Implement a Salmonella monitoring program, if applicable, meeting the requirements
stated in applicable husbandry quality systems (e.g. IKB, KKM, QM-Milch). In case of
home mixing of feed or drying of feed, the monitoring program on feed in Appendix XII is
referred to. Concerning Salmonella monitoring programs for the transport and storage of
feed is referred to appendices XI and XI respectively. For specific information concerning
Salmonella monitoring in husbandry, see below. On the products or places, which were
determined of risk by the HACCP study (e.g. in feeding systems, troughs (incl. in milking
parlour), buckets, teats, machinery used for feeding or mixing feed, water, silos, milking
equipment, milk tank and feed of risk as mentioned in chapter 8);
Implement a Salmonella monitoring program on products which were determined of risk
by the HACCP study: e.g. feed (incl. on farm produced products of vegetal origin and on
farm produced dairy products (e.g. raw milk)), compound feed, silages, milk replacers,
additives, premixtures, water (e.g. drinking water, well water), residues and spills in
specific areas (e.g. in trench silo, in feeding system), manure, raw milk and eggs;
Implement a Salmonella monitoring at critical points in husbandry, as is determined using
the HACCP system. Also include machinery and equipment in the monitoring program. A
representative number of samples should be taken and examined from the critical points.
Critical points where samples can be taken are e.g.:
o Dust from ledges inside and the top of feed silos / bins;
o Feeding systems;
o Troughs (incl. milking parlour);
o Buckets and teats (e.g. used for milk replacer feeding);
o Machinery used for mixing feed or other machinery in direct contact with feed;
o Milking equipment;
o Equipment in direct contact with feed;

APPENDIX X Salmonella control measures: transport of feed.
be complied with (GMP+3, 2012). Specific requirements and control measures related to
transport are stated in various GMP+ standards (GMP+6,7,8,9,10,11, 2012). The applicable
GMP+ standard depends on the means of transport. The applicable standard should be
complied with. Additional recommendations for general control measures are stated below in
this appendix.
Transport
Use the HACCP system to assess the risk of Salmonella in the feed during transport.
Besides others, the following items should be addressed:
Mind the temperature of feed at loading. The temperature should be low enough to
ensure that no condensation and heating will occur;
If covers are used during transport, they must be maintained in a clean condition by
being cleaned, sanitised and dried regularly (EFSA6, 2008);
If transport can be made airtight, fumigate if pests are present (FAO2, 2007).
Salmonella monitoring programs: transport

2012). Several types of monitoring programs are suggested: on feed and on critical points in
the storage of feed, as stated below:
Implement a Salmonella monitoring program on feed that is determined of risk by the
HACCP study, also residues and spills in specific areas;
Implement a Salmonella monitoring program inline at critical points in transporting feed,
as is determined using the HACCP system. Also include machinery and processing
equipment in the monitoring program. A representative number of samples should be
taken and examined from the critical points. Critical points where samples can be taken
are e.g.:
o Surfaces of e.g. trailers, containers and holds;
o Covers;
o Equipment used during loading and unloading;
o Dust bag.
In the event of a positive finding analysis the Salmonella monitoring frequency should
increase. The positive samples should be classified.

APPENDIX XI Salmonella control measures: storage of feed.
be complied with (GMP+3, 2012). Specific requirements and control measures related to
storage are stated in the GMP+ B5 standard (GMP+12, 2012). Additional recommendations
for general control measures are stated below in this appendix.
Storage
Use the HACCP system to assess the risk of Salmonella in the feed during storage and
transhipment. Besides others, the following items should be addressed:
Drying of feed, if the feed is too moist. Dry up to a moist content unfavourable for the
growth of Salmonella. The dried feed should be cooled up to a temperature as low to
prevent condensation and heating during storage;
A pest control program should be in place: see section Pest control;
Make a continuing effort to minimize storage time of feed, thus decreasing the
opportunity for contamination with Salmonella and other microbes (AFIA, 2010).
Salmonella monitoring programs: storage

2012). Several types of monitoring programs are suggested: on feed and on critical points in
the storage of feed, as stated below:
Implement a Salmonella monitoring program on feed that is determined of risk by the
HACCP study, also residues and spills in specific. The minimum monitoring requirements
per feed for GMP+ certified participants are stated in GMP+ Appendix BA4 (GMP+1,
2012);
Implement a Salmonella monitoring program inline at critical points in storing feed, as is
determined using the HACCP system. Also include machinery and processing equipment
in the monitoring program. A representative number of samples should be taken and
examined from the critical points. Critical points, besides the ones indicated in the GMP+
standards, where samples can be taken are e.g.:
o The unloading pit / dumping pit for feed (SVA, 2011), e.g. dust from auger system
(DEFRA, 2009);
o Dust from ledges inside of ingredient silos / bins, or dust from sieves (DEFRA, 2009);
o Dust from air aspiration cyclone system. Unless this is not accessible vacuumed or
swept dust could be used (DEFRA, 2009);
o Dust from vacuum cleaners (DEFRA, 2009);
o Dust from augers, conveyer belts, elevators;
In the event of a positive finding analysis the Salmonella monitoring frequency should
increase. The positive samples should be classified.

APPENDIX XII Salmonella control measures: processing of feed.
In GMP+ B1 and B2 standard general control measures and requirements are stated which
should be complied with (GMP+3,13, 2012). Additional recommendations for general control
measures are stated below in this appendix.
Purchasing and receiving feed
Verify if the supplier / producer of feed (also include premixes, additives etc.) has
assessed the hazard and risk of Salmonella;
Selecting of the suppliers based on their implementation of programs for the control of
Salmonella (EFSA6, 2008) and hygiene;
All feed arriving at the production site should be subjected to arrival inspections and
include the following (AFIA, 2010):
o Inspection of documentation, invoices and seals (if applicable);
o Assessment of transport entities with respect to maintenance, sanitation, cleanliness
and, if applicable, covered transport entities;
o Verification of ingredient identity (e.g. by documents, visual inspection);
o Inspection of ingredient for quality indicators, including the following:
Visible evidence of water damage;
Visible faecal or vermin contamination;
Temperature check;
Aroma;
o Procedures for dealing with Salmonella risk feed and Salmonella-positive feed (e.g.
quarantine);
The receiving area should be designed for wet weather operation, so that the unloading
of ingredients occurs without significant water damage to the ingredients (AFIA, 2010).
Conditioning, heat or other technological treatment (e.g. pelleting, expanding, extrusion)
The use of high temperatures to accomplish pasteurisation during processing is based on the
destructive effects of time and temperature on Salmonella and other microorganisms.
Microbiologists have identified at least 11 factors or parameters of microorganisms and their
environment that can affect heat destruction. These factors include moisture or water activity,
fat levels, presence of salts, presence of carbohydrates, pH, protein content, number of
organisms, age of organisms, inhibitory compounds, and time and temperature history.
Despite the influence these factors can have on the resistance of microorganisms to heat,
thermal destruction during the processing steps of pelleting or extrusion is the most critical
control step for destruction or reduction of Salmonella and other pathogenic microorganisms
(AFIA, 2010).
Microbial populations are not killed instantly upon exposure to heat, moisture and
pressure. One second of moist heat (at 22% moisture and 106 log initial population) at
77C can kill Salmonella organisms as long as all processed material actually reaches
these recommended temperatures internally or throughout. While this should be the
target or intended minimum processing temperature, it may not be reached during the
start-up phase of the pelleting or extrusion process. For this reason, it may be necessary
to recycle the first material coming through the pelleting or extrusion process to allow for
system warm-up and adequate processing temperature (AFIA, 2010);
The effective heat treatment should be determined at the individual plants. Although both
time and temperature are crucial in determining the number of Salmonella destroyed by
the pelleting process, several researchers have suggested that target temperature of 80
to 85C be established for conditioning (Jones and Richardson, 2004). Conditioning
times in conventional equipment generally range from a few seconds to several minutes,
depending on the equipment involved. The pelleting process can also be augmented with
the use of expanders (Jones, 2011). The EFSA6 (2008) also includes conditioning prior to

Conditioning, heat or other technological treatment (e.g. pelleting, expanding, extrusion)
pelleting or expanding as usual processing procedures performed in most feed mills

when heat treated feed is manufactured. Extended steam conditioning up to 30 seconds
or more have been shown to increase the antibacterial effect of the heat treatment. For
these reasons it is therefore important that pellet mills are carefully monitored to insure
proper operation with respect to temperature and retention time in the conditioner
particularly the period after the production started when optimum antibacterial conditions
do not prevail (EFSA6, 2008). Lower temperatures can also be affective however a longer
heating time should be applied (OVOCOM2, 2008).
Decreasing the pH will reduce Salmonella contamination. This can be achieved by
various methods (OVOCOM2, 20008):
Method Maximum pH
Lactic acid fermentation 4.5
Addition of organic acids 4
Addition of inorganic acids 3.5
Although heat treatment is generally recognised as the most effective decontamination

method, in some circumstances (e.g. pelleted feed for layers) this may not be
appropriate. In such cases, chemical treatment of feed may offer an alternative means of
protection (EFSA6, 2008). Other methods should be validated and verified by
experimental studies (OVOCOM2, 2008).
The effectiveness of the treatment in killing Salmonella should be verified.
After heat treatment, cooling and drying should occur as rapidly as possible to reduce
condensation in processing and conveying equipment, storage containers, packaging
equipment and in the distribution of finished products (AFIA, 2010);
Thermocouples or temperature monitoring devices should be strategically placed in the
process flow to monitor the actual temperature (AFIA, 2010);
Coolers and dryers
After heat processing, cooling and drying should occur as rapidly as possible to reduce
condensation in processing and conveying equipment, storage entities, packaging
equipment and in the distribution of finished products (AFIA, 2010);
Critical points concerning Salmonella recontamination have been defined in coolers.
Packaging
In case of bagging, new bags (incl. big bags) are recommended for finished product
packaging. Previously used bags should be sanitised before used for packaging (AFIA,
2010).
Salmonella monitoring programs: processing

2012). Three types of monitoring programs are suggested:
Implement a Salmonella monitoring program on the following ingredients and
intermediate products, which were determined of risk by the HACCP study: e.g. feed,
processing aids, water, pressed meal before and after heat treatment, residues and spills
in specific areas. Special attention should be paid to Salmonella critical feed. The

Salmonella monitoring programs: processing
minimum monitoring requirements per feed for GMP+ certified participants is stated in
GMP+ Appendix BA4 (GMP+1, 2012);
Implement a Salmonella monitoring program inline at critical points in processing feed, as
is determined using the HACCP system. Also include machinery and processing
equipment in the monitoring program. A representative number of samples should be
taken and examined from the critical points. Critical points, besides the ones indicated in
the GMP+ standards, where samples can be taken are e.g.:
o The unloading pit / dumping pit for feed (SVA, 2011), e.g. dust from auger system
(DEFRA, 2009);
o Dust from ledges inside of feed silos / bins, or dust from sieves (DEFRA, 2009);
o Dust from hammer mills (DEFRA, 2009);
o Inside of coolers where condensation sites are possible e.g. the top of the pellet
cooler (SVA, 2011);
o Dust from coolers, taken below coolers or on associated framework, ledges etc.
(DEFRA, 2009);
o Dust from air aspiration cyclone system. Unless this is not accessible vacuumed or
swept dust could be used (DEFRA, 2009);
o Dust from vacuum cleaners (DEFRA, 2009);
o Dust from pellet shakers (DEFRA, 2009);
o Dust from augers, conveyer belts, elevators;
Implement a Salmonella monitoring program on feed (e.g. compound feed, feed
ingredients directly delivered to farmers (e.g. cereals)), with a higher frequency of
analysis in feed of high(er) risk (based on the HACCP study). The minimum monitoring
requirements per feed delivered directly to farmers for GMP+ certified participants are
stated in GMP+ Appendix BA4 (GMP+1, 2012);


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Fact sheet:

GMP+ International B.V.

All rights reserved. The information in this publication may

DISCLAIMER: In spite of all the care and attention given to

Summary of GMP+ products standards for the animal feed sector

Fact sheet: Salmonella 2 / 57

Fact sheet: Salmonella 3 / 57

Fact sheet: Salmonella 4 / 57

Fact sheet: Salmonella 6 / 57

- Consumption chicks - 0+% 20(approaching

Laying poultry feed: end products and feed GMP+

- Raising increase laying poultry - 0+% 20(approaching

- Laying-hens and breeding hens 1% 0+% 20(approaching

- Raising increase turkeys - 0+% 20(approaching

- Increase turkeys - 0+% 20(approaching

Fact sheet: Salmonella 7 / 57

- Spontaneous lactic acid fermentation - 4.5 These norms do not apply

Fact sheet: Salmonella 8 / 57

This fact sheet focuses on Enterobacteriaceae and Salmonella contamination of feed1 as a

1. Nature, history and prevalence of Enterobacteriaceae and Salmonella

Fact sheet: Salmonella 9 / 57

Table 2. CDC Salmonella nomenclature (Brenner et al., 2000).

Fact sheet: Salmonella 10 / 57

A microscopic picture of Salmonella is shown in figure 1.

Figure 1. Salmonella (red colour) (source: Wikipedia)

Depending on the serotype the effect of different environmental circumstances of Salmonella

Salmonella has long been recognised as an important zoonotic pathogen of economic

Fact sheet: Salmonella 11 / 57

Fact sheet: Salmonella 12 / 57

Other sources than feed ingredients of Salmonella contamination of animal feed:

Fact sheet: Salmonella 13 / 57

Fact sheet: Salmonella 14 / 57

Fact sheet: Salmonella 15 / 57

Fact sheet: Salmonella 16 / 57

Fact sheet: Salmonella 17 / 57

In the DOS database data can be found concerning Salmonella in feed.

Fact sheet: Salmonella 18 / 57

Fact sheet: Salmonella 19 / 57

Fact sheet: Salmonella 20 / 57

Fact sheet: Salmonella 21 / 57

4. Potential adverse effects

Fact sheet: Salmonella 22 / 57

In Appendix I the potential adverse effects of Salmonella in animals are shown.

In Appendix I the potential adverse effects of Salmonella in humans are shown.

5. Severity of the potential adverse effects

The severity of Salmonella toxicosis in animals is classified as high because:

The severity of Salmonella toxicosis in humans is classified as high because:

Table 7. Severity of Salmonella toxicosis

Fact sheet: Salmonella 23 / 57

Fact sheet: Salmonella 24 / 57

Table 8. Products with a higher risk to contain Salmonella.

and their excrements of vermin or manure;

Products from the oil crushing industry:

and expeller, soy protein concentrate, toasted soy beans);

Fact sheet: Salmonella 25 / 57

Fact sheet: Salmonella 26 / 57

Fact sheet: Salmonella 27 / 57

Fact sheet: Salmonella 28 / 57

Fact sheet: Salmonella 29 / 57

Fact sheet: Salmonella 30 / 57

Fact sheet: Salmonella 31 / 57

Fact sheet: Salmonella 32 / 57

Fact sheet: Salmonella 33 / 57