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1
Microbial Risk Assessment Project
‘What is the risk of infection from Salmonella spp. in the Republic of Ireland from eating ready-to-eat
fruit and vegetables?’
Francis Higgins, Cliodhna McMahon and Karen Duffy
Department of Microbiology
National University of Ireland, Galway
March 2015
2
Contents
1. Executive Summary: Pg. 3
2. Statement of Purpose: Pg. 4
3. Hazard identification: Pg. 4
4. Hazard characterisation: Pg. 5
5. Exposure Assessment: Pg. 16
6. Risk characterisation: Pg. 27
7. Discussion: Pg.
8. Conclusions: Pg.
9. Bibliography: Pg.
4
2. Statement of Purpose
The aim of this project is to identify the risk of Salmonella infections from consumption of
ready-to-eat fruit and vegetables in the Republic of Ireland through qualitative analysis of
each step undertaken by the products during their journey from ‘farm to table’. Extensive
research was carried out using journals, scientific papers, national health and food
publications and online resources in order to determine the probable level of risk associated
with each step.
3. Hazard identification
Salmonella spp. are a group of rod shaped, oxidase negative, gram-negative facultative
anaerobes belonging to the family Enterobacteriaceae (Christenson, 2013).
Salmonella precedes Campylobacter spp. as the cause of most zoonotic infections in Europe
(Jordan et al, 2006). It is located in the intestinal tract of animals and humans, and is spread
through faeces. There is an average of 350-450 cases of salmonellosis annually in the
Republic of Ireland (HPSC, 2009). Contaminated food is regarded as the number one mode
of transmission of Salmonella spp. into humans, leading to salmonellosis, costing the health
care system significant sums of money annually (FSAI, 2011). Salmonella spp. have also
been known to be transmitted from animals to humans, and humans to humans (D’aoust,
2000).
There are only 2 species of Salmonella - S. enterica and S. bongori. However, more than
2400 Salmonella serotypes have been identified (Adak et al, 2005). They are facultative
anaerobes and are oxidase negative. They can grow at anywhere between 5 - 46ºC, but grow
optimally at 35ºC - 45ºC. They can survive for extended periods of time in refrigeration
(shown to survive for 28 days on surface of vegetables stored in fridges). Antimicrobial
strains are becoming more prevalent around the world which is an increasing cause of
concern.
An infective dose of Salmonella spp. (i.e., the amount of bacteria needed to be ingested to
cause illness) can be as low as 5-20 cells (FDA, 2012). Salmonellosis typically manifests as a
self-limiting case of enterocolitis. This is characterized by abdominal pain, diarrhoea, fever,
nausea, vomiting and chills. The incubation time is 5 – 72 hours with recovery time ranging
5
from 2 – 5 days depending on the strain and host characteristics (D’aoust, 2000). 93% of
individuals showing symptoms of salmonellosis recover without seeking medical attention.
Typically no treatment is needed other than taking oral fluids. In severe cases of dehydration
fluids may need to be replenished through an intravenous drip (CDC, 2013). Complications
may arise leading to systematic infections, and can also be a cause of various chronic
conditions, e.g., reactive arthritis (FSAI, 2011). All groups are susceptible to salmonellosis,
with the highest number of incidences being seeing in children 0 – 4 years old. In adults, it is
estimated that 1-3 working days are lost due to an occurrence of the illness (D’aoust, 2000).
Salmonella spp. are prevalent in many animals, both domestic and wild. They can pass
through the food chain all the way to homes/ food catering services. Humans generally
contract salmonellosis through consuming contaminated foods, including those of animal
origin (meat, eggs, poultry and milk) and vegetables and fruit that become contaminated, e.g.,
from manure or irrigation water (WHO, 2013). Imported fruit and vegetables that are
irrigated with untreated wastewater are of particular concern (Ait Melloul et al, 2001).
Non-typhoid salmonellosis is a notifiable disease in the Republic of Ireland. Data obtained
from the Food Safety Authority of Ireland (FSAI) showed that 333 cases of salmonellosis
were reported in 2009 (a crude incidence rate of 7.9 cases per 100,000) (FSAI, 2011). This
was a part of a large outbreak in the EU, which totalled 108,614 human cases of
salmonellosis. A crude incidence rate of 10.6 per 100,000 was seen in Ireland the previous
year. Due to the ability of Salmonella spp. to cause illness at considerable low doses, it is of
the upmost importance to protect consumer health through vigilant controls at all stages of
preparation and supply of foods.
4. Hazard characterisation
There are numerous host factors that have an effect on the pathogenicity of Salmonella spp.
following ingestion, including (but not limited to): age, gender, nutritional status, immune
status and previous exposure (WHO/FAO, 2002). These factors cause a variation in the
infective dose of the bacteria, which may be very high (>10 5 cells) or in the 10s of cells
(Hara-Kudo & Takatori, 2011).
6
It is a common trend that the highest numbers of incidences of reported Salmonella infections
are found in young children and elderly people (WHO/FAO, 2002). Although the majority of
these cases are usually mild, children and the elderly also possess a higher occurrence of the
infection progressing to a more severe state than with other age groups, including death in
some instances. However, the associations of these two age groups with the illness may have
an effect on data pertaining to their increased incidence rates. For example, medical care is
more likely to be sought after for infants and elderly showing symptoms relating to
salmonellosis infections than in adults (and subsequent testing for Salmonella infections).
Furthermore, young children are far more susceptible to infection if it is their first incidence
of exposure to the bacteria (WHO/FAO, 2002).
The immune status of the host is also a considerable factor in the occurrence of infection
from Salmonella spp, as it is with any other disease causing bacteria. People who are
immunocompromised or suffer from debilitating illnesses are particularly susceptible to
developing salmonellosis from eating contaminated food, such as fruit and vegetables, and
are also at greater risk at repeat infections or progression of the infection to more severe
ailments (WHO/FAO, 2002). In healthy people, the development of immunity has been seen,
whereby patients who were voluntarily infected with S. enterica twice had less severe
symptoms from the second trial, even though they were administered a higher dose of the
bacteria the second time round (McCullough and Eisele, 1951). Further evidence suggests
that immunity to Salmonella is serotype specific due to the higher incidences of salmonellosis
in people who are travelling and thus exposed to new serotypes, often from the consumption
of fruit and vegetables in foreign countries (WHO/FAO, 2002).
Virulence plasmids have been associated with the pathogenicity of different Salmonella
strains and their ability to spread after colonizing the stomach, invade the intestine,
proliferate in the spleen and suppress the immune responses of their hosts (Slauch, Taylor &
Maloy, 1997). A study investigating the presence of virulence plasmids in different strains of
Salmonella found that all isolates of the highly invasive serotypes S. Enteritidis, S. Dublin
and S. Choleraesuis.
The clinical manifestations of salmonellosis range from general gastroenteritis symptoms to
enteric fevers (e.g. typhoid fever) (Giannella, 1996). The incubation time for Salmonella
gastroenteritis is dependent on the dose that is ingested. Initial sympoms include vomiting,
nausea, diarrhoea and abdominal cramps (with myalgia, headache, fever and chills also
7
arising in many cases). Enteric fevers are life threatening systematic sicknesses which require
swift antibiotic treatment. The most comprehensively studied of these is typhoid fever
(caused by S. Typhi). The incubation period for enteric fevers caused by salmonellae is
typically 10 – 14 days, with gastroenteritis often preceding it. Other syndromes arising from
salmonellosis include an asymptomatic carrier state, focal infections and septicaemia.
Although any of the serotypes can be responsible for any of these syndromes arising, some
serovars have been strongly linked to particular syndromes, for example:
Enteric fever – S. Typhi, S. Paratyphi, S. Schottmuelleri
Septicemia/ focal infections – S. Choleraesuis
Gastroenteritis – S. Typhimurium, S Enteritidis
The majority of non-typhoidal salmonellae are ingested in contaminated food. Salmonellae
must have a number of virulence factors in order to be pathogenic, including:
Ability to invade cells
Lipopolysaccharide coat
Ability to replicate in cells
Production of toxins (in some cases)
Once ingested, the ilium and colon are colonized by the bacteria (Giannella, 1996). They then
invade the epithelial cells of the intestines (and the lymphoid follicles) where they proliferate.
When invading the epithelial cells the bacteria first bind to specific receptors found on the
cell surface. They then induce ruffling of the enterocyte membrane, followed by pinocytosis
of the bacterial cells. Multiple genes (located on chromosomes and plasmids) are involved the
attachment and invasion mechanisms of the pathogens into host cells.
In order to diagnose salmonellosis, bacterial isolates are required. (Giannella, 1996)
Biochemical tests are used to identify the genus Salmonella while serological testing is used
to confirm the serological type. Samples (typically faeces or blood) are then plated on
selective and non-selective agars, as well as enrichment broths.
There are vaccines for treating typhoid fever that are moderately effective (particularly in
children) (Giannella, 1996). However no vaccines currently exist for treating non-0thyphoidal
salmonellosis, although research is on-going for their development. To treat general
salmonellosis oral and intravenous replacement of lost fluid is implemented, along with
8
medication to control and nausea, vomiting or pain. Antibiotics are administered for typhoid
fever and enteric fever, while with non-typhoidal salmonellosis they are only used for cases
of septicaemia, focal infections syndromes and enteric fever. They are typically not used for
normal gastroenteritis symptoms as they do not reduce the duration of the illness, but do
extend the period of faecal excretion of the pathogens and run the risk of producing more
antibiotic-resistant strains of the bacteria.
Epidemiology of Salmonella spp. infections in the Republic of Ireland and Europe
Figure 1: Reported no. of confirmed S. Agona cases, sorted by age and gender, S. Agona
Outbreak, Europe, Summer 2008 (N=163). (NSRL, 2008)
163 confirmed cases of Salmonella were reported between February and September 2008
across Europe. These cases affected a range of people differing in ages (NSRL, 2008). The
youngest affected was 3 months old, to the oldest affected who was 87 years old. The median
age affected was 27 years. 57% of the people affected were males. During this outbreak 25
people were hospitalised, and two elderly patients died. The first reported dead was a 77 year
old female who contracted Salmonella whilst in hospital, the second dead was a 78 year old
male. Both cases occurred in the UK.
9
Figure 2: Distribution of confirmed S. Agona cases, S Agona Outbreak, Ireland and the
UK, Summer 2008 (NSRL, 2011)
Eleven cases were reported in Ireland. England reported the majority of cases - 96. Thirty
four cases were identified in Scotland and 11 in Wales. Two cases were reported in Northern
Ireland.
Figure 3: Epidemic Curve - Reported number of confirmed cases by date of onset and
country of infection, S. Agona Outbreak, Europe, Summer 2008 (N=163). (NSRL, 2011)
10
During the S. Agona outbreak in 2008, 56 cases were interviewed to identify the source of the
infection. 40 cases stated that they ate takeout food and 38 people said that they had
consumed made-to-order sandwiches before becoming ill. The fillings in the made-to-order
sandwiches were unknown at the time of the study.
Figure 4: Seasonal variation of Salmonella isolates (NSSLRL, 2010)
Figure 5: Number of cases and CIR of human salmonellosis in Ireland, 2008 (HPSC,
2010)
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Figure 6: Number of cases of salmonellosis in Ireland for years 1990-2008 (HPSC, 2010)
Salmonellosis cases in Ireland have remained reasonably constant in the last eight years.
There are approximately 350-450 cases confirmed each year (HPSC, 2010). The cases of
salmonellosis have decreased from the late 1990’s, where the incidences of cases confirmed
rose to 1257 cases in 1998. 26% of the confirmed cases in 2008 were found to be children
under the age of 5. The age specific incident rate was 46 per 100’000 in this age group. This
was similar across all HSE-areas apart from the North West which showed the highest
incidence was in the 20-24 year old group. Salmonella was isolated from 447 people. These
isolates were referred to the National Salmonella Reference Laboratory for further typing; it
was found that 12 of the isolates were S. typhi or S paratyphi. The remaining isolates were
non-typhoidal Salmonella. S. Enteritidis (28%) and S. Typhimurium (32%) were the most
common serotypes detected. The number of cases of S. Typhimurium exceeded the number
of cases of S. Enteritidis. S. Agona with found in 13 isolates, most of these were found in the
international outbreak in 2008.
12
Figure 7 : Non-typhoidal Salmonella isolates referred to NSRL by serotype and HSE
area, Ireland 2008 (HPSC, 2010)
Figure 8: Salmonella serotypes reported on CIDR by age groups 2008. (HPSC, 2008)
13
Figure 9: Clinical isolates of S.enterica (n=444) sorted by age and gender from NSRL 2007 (HPSC, 2007)
456 cases of salmonellosis were confirmed in Ireland in 2007 (HPSC, 2007). 440 of these
cases were confirmed in a laboratory. All possible cases were linked to Salmonella outbreaks
on CIDR. 457 cases of Salmonella isolates were referred to the National Salmonella
Reference Laboratory. In 2007 there was a slight predominance in males. The female: male
ratio in 2007 was 1:1.1. The highest number of cases occurred in children under five years of
age. 24% of cases occurred in this age category
Figure 10: Incidence rate of salmonellosis in Ireland 2007 by age group (HPSC, 2007)
14
The national crude incidence rate of salmonellosis cases for 2006 was 10.0/100’000. In 2007
there was an increase observed to 10.7/100’000. Most significantly the incident rate of
salmonellosis in the HSE‐S region increased from 8.3 in 2006 to 21.1 in 2007. This may be
due to an outbreak of S Enteritidis. 52 cases occurred in HSE-S
Figure 11 & 12: Number of cases [2006 and 2007] and CIR of human salmonellosis
[’00-‘07] in Ireland (HPSC, 2007)
It was stated that the CIR of salmonellosis cases were similar in 2006 and 2007 (HSPC,
2007). There was further investigations into regional rates and found that the cases were not
similar. The CIR in the regions of HSE‐M, HSE‐NW and HSE‐S experienced a change in
rate of greater than one case per 100,000. In the midland and the North West a decline in
cases occurred, from 14.3 to 19.1 per 100,000 in the midlands and 16.0 to 8.9 per 100,000 in
the north‐west. The CIR of the southern region increased from 8.3 per 100,000 to 21.1 per
100,000 this increase is probably due to an outbreak of S. Enteritidis. The CIR in Northern
Ireland was slightly lower with an estimated rate of 9.0 per 100,000 likely for the year. Cases
of Salmonellosis were observed in all age groups. There was a higher incidence in the 0‐4 age
bracket. This is probable due to clinicians seeking clinical samples in children under five.
15
The Number of male and female cases has been similar in past years, female: male ratio of
1:1 in 2007, 1.3:1.0 in 2006 and 1:0.9 in 2005 (HSPC, 2007). Serotyping data showed that S
Typhimurium and S Enteritidis remain the most prominent strains. 64 serotypes were
identified by the NRSL in 2007. The number of S. Typhi and S. Paratyphi A cases in 2007
increased from previous years. There were 9 cases of S. Typhi in 2003, five cases in both
2004 and 2005, and, seven in 2006. There was only one case of S. Paratyphi A in 2006.
A Review of European data shows the rate of Salmonella is decreasing in continuing in
2007. Salmonellosis is still the second most commonly reported zoonotic disease in Europe.
The major sources of human Salmonella infections are due to the consumption of eggs, pork
and chicken. There were 151,995 confirmed cases notified on the European Surveillance
System in 2007. This is 31.1 cases per 100,000. CIR’s varied by country by as much as 2.9
confirmed cases per 100,000 to 171.6 confirmed cases per 100,000. S. Enteritidis and S.
Typhimurium were the most common serovars reported they accounted for 81% of all known
serovars in human cases.
The most dominant serotypes of Salmonella in Ireland are S. Enteritidis and S. Typhimurium
(McKeown et al. 2012). These two serotypes accounted for 20% and 38% respectively, of
human clinical isolates identified in Ireland in 2010, while other serotypes made up the
remaining 42% of isolates. DT104 and DT104b have been the most common phage types
detected. In Ireland overseas travel plays an important role in Salmonella epidemiology. It is
now estimated that up to half of all notified cases may be travel associated.
17
5. Exposure assessment
Table 1: Variables and data inputs for the exposure pathway for Salmonella spp. in
ready-to-eat fruit and vegetables (P=Probability) (C=Concentration of m/o)
Main Stage Sub Stage Outputs Inputs
Farm Seed/ Plant
imports
P1: The probability that the
imported seeds/plants are
carrying Salmonella.
C1: The concentration of
Salmonella in the imported
seeds/plants
In the production of seeds, the
practice of animal grazing to
initiate flowering can result in
the introduction of enteric
bacteria in faeces.
Allowing wild or domestic
animals to graze in seed fields
can also cause contamination
of seeds.
Regional difference
Season of infection
Management factors
Seed/ Plant
transport
vendors
P2: The probability that the
seeds/plants will be exposed to
Salmonella during transportation
C2: The concentration of
Salmonella contacted by the
seeds/plants during
transportation
P1,C1
Conditions of the containers
used for the transport the
seeds?
Storage conditions in vehicles
used for transport?
What is the duration of the
transport journey?
Avian P3: The probability that birds
defecating causes the soil to be
contaminated with Salmonella
C3: The concentration of
Salmonella in bird faeces
P2,C2
Birds defecating onto soil,
plants, fertiliser, irrigation
water or manure can cause
contamination of the produce
18
with Salmonella
Growth/survival dynamics
Manure/
irrigation water/
fertilizer
P4: The probability that the
manure/irrigation/fertiliser
spread onto soil is contaminated
with Salmonella
C4: The concentration of
Salmonella present in the
manure/fertiliser/irrigation water
P3,C3
Improperly treated manure
can contain Salmonella.
Irrigation water or fertiliser
can become contaminated
with Salmonella due to wild
or domestic animals
defecating into these sources.
Growth/survival dynamics
Management factors
Insects P5: The probability that insects
are carrying Salmonella and can
contaminate the produce plants
C5: The concentration of
Salmonella present in the insects
P4,C4
Insects can be carriers of
Salmonella onto the
plant/produce.
Insect defecate onto plants
during feeding and
contaminate plants with
Salmonella.
Soil P6: The probability that the soil
is contaminated with Salmonella
C6: The concentration of
Salmonella in the soil
P5,C5
Soil can carry pathogenic
bacteria. Spraying of soil with
improperly treated manure can
cause contamination
Wild or domestic animals
defecating onto soil can lead
to the transmission of
Salmonella from the host to
the soil.
Survival/growth dynamics
Harvesting of
fruit and veg
P7: The probability that the
fruits and vegetables will be
P6,C6
Harvesting equipment may be
19
contaminated during harvesting
C7: The concentration of
Salmonella contracted by the
vegetables during harvesting
contaminated with pathogens
and can cause the fruit and
veg to become contaminated
Management factors
Survival dynamics
Washing of
produce
P8: The probability of the fruit
and vegetables being exposed to
Salmonella during washing.
C8: The concentration of
Salmonella present in the wash
water
P7,C7
Improperly washed fruit and
vegetables may still contain
Salmonella
Temperature of the water used
for washing
How clean the water is for
washing?
Management factors
Storage on Farm P9: The probability of the fruit
and vegetables contracting
Salmonella during storage on the
farm
C9: The concentration of the
pathogen being contracted
during storage on the farm
P8,C8
The storage containers on the
farm may not be cleaned
properly and can cause
contamination
Temperature and duration of
storage and how these may
vary
Packaging on
farm
P10: The probability that the
fruit and vegetables with be
exposed to Salmonella during
packaging on the farm
C10: The concentration of
Salmonella being contracted
during packaging
P9,C9
Human handling can cause
contamination
Faecal matter spread from
humans to produce
Other sources of
contamination
20
Distribution &
Storage
Transport of
fruit and veg
P11: The probability of the fruit
and vegetables being exposed to
Salmonella during transport
C11: The concentration of
Salmonella present in the
transporting environment
P10,C10
Conditions of the containers
used for the transport the
produce?
Storage conditions in vehicles
used for transport?
What is the duration of the
transport journey?
Wholesale
market
P12: The probability of the fruit
and vegetables becoming
exposed to Salmonella in the
wholesale market
C12: The concentration of
Salmonella present in the
wholesale market environment
P11,C11
Improper handling from
sellers/customers examining
produce.
Contaminated containers
Storage conditions in market
Duration of produce at
wholesale market
Management factors
Processing
station
P13: The probability that the
fruit and vegetables will contract
Salmonella during processing,
such as cutting, slicing and
dicing
C13: The concentration of
Salmonella that could be
contracted by the produce in the
processing station
P12,C12
Contaminated surfaces
Contaminated equipment
Improper handling of produce
Other sources of
contamination
Preliminary
washing
P14: The probability of the
produce becoming contaminated
with Salmonella during washing
with chlorinated water
C14: The concentration of the
pathogen present in chlorinated
water
P13,C13
Washing produce with
unclean water
Human handling can
contaminate produce after
washing
Temperature of the water used
21
for washing
Slicing or
shredding
P15: The probability of the
produce coming into contact
with the pathogen during slicing
and shredding
C15: The concentration of the
pathogen present on the slicing
and shredding equipment
P14,C14
Unclean equipment
Cross contamination between
equipment and produce
Unclean surfaces
Washing/
disinfection
P16: The probability of the
produce becoming contaminated
with Salmonella during
disinfection
P15,C15
Improperly washing the
produce
Improperly disinfection of
produce
Temperature of water used?
How clean is the water that is
used for disinfection?
Growth/survival dynamics
Packaging P17: The probability that the
fruit and vegetables with be
exposed to Salmonella during
packaging
C17: The concentration of
Salmonella being contracted
during packaging.
P16,C16
Is the package contaminated?
Damage to the packaging can
cause contamination of
produce
Improperly sealed packaging
Human handling
Retail & Home Retail storage P18: The probability of the fruit
and vegetables being exposed to
Salmonella during storage in
retail facilities
C18: The concentration of
Salmonella present in storage
containers in retail facilities
P17,C17
Storage time and temperature
in retail facilities
Produces stored in clean
containers
Human handling of produce
Management factors
Survival dynamics
Other sources of
22
contamination
Consumer
transport home
P19: The probability of the fruit
and vegetables being exposed to
Salmonella during transportation
to consumers home.
C19: The concentration of
Salmonella present in the
transporting environment
P18,C18
Conditions of the containers
used for the transport the
produce?
Storage conditions in vehicle
used for transport?
What is the duration of the
transport journey?
Temperature during transport?
Storage in home P20: The probability of the
produce becoming contaminated
with Salmonella in the
consumer’s home.
C20: The concentration of
Salmonella contracted by the
produce whilst present in the
consumers home
P19,C19
Storage time and temperature
in the home and how these
vary?
Poor hygiene in home
Potential cross-contamination
Survival/growth dynamics
Human handling
Prep before use P21: The probability of the fruit
and vegetables becoming
contaminated with Salmonella
during preparation
C21: The concentration of
Salmonella present on the fruit
and vegetables during
preparation
P20,C20
Unclean surfaces
Unclean equipment
Improperly washing of
produce
Cross contamination from
other sources
23
Serving size S: Serving size Amount of produce consumed
in any serving and variation of
this within and between
different population
groups( ages, sex, regions,
health)
Estimate of
exposure
P: Probability of exposure of
Salmonella per serving
C: Number of organisms
consumed per serving
P21,C21,S
Date relating to stages of exposure pathway of Salmonella spp. in ready-to-eat fruit and
veg:
Pathogens can contaminate fruit and vegetables during different points throughout pre-
harvesting and post-harvesting systems. (Beuchat, 2002) Potential pre-harvest sources of
microorganisms include soil, faeces, irrigation water, dust, insects, manure, wild and
domestic animals, and human handling. Post-harvest sources include faeces, human handling,
harvesting equipment, transport containers, wild and domestic animals, insects, dust, rinse
water, ice, transport vehicles, and processing equipment. Containers that are used for harvest,
24
transport, and display raw fruits and vegetables may not be cleaned and sanitized, which can
lead to the development of biofilms.
Before harvest:
Most microorganisms are naturally present in the environment. These microorganisms can be
non-pathogenic or pathogenic. Pathogenic organisms can contaminate fruit and vegetables
throughout many stages from farm to fork. Pre-harvest contamination can occur by animal
faeces by wild or domestic animals, dust, irrigation waters, runoff water, and insects. They
may also be contaminated from septic tanks leaching through soil.
Microbiological analyses have found that alfalfa seeds can contain high levels of
microorganisms, which include faecal coliforms (NACMCF, 1999).
The environmental conditions under which seeds are sprouted (such as growing time,
temperature, moisture and nutrients) are ideal for bacterial proliferation. It is found that an
increase of 100 to 1000 fold of microorganisms can occur on the sprouted seeds without
affecting the appearance of the product (Taormina et al, 1999).
Several pathogens have been implicated in sprouted seed-associated outbreaks these include
Salmonella, enterohaemorrhagic E. coli, Bacillus cereus, and Listeria monocytogenes. (López
Camelo, 2002)
Salmonella in soil:
Salmonella is a microorganism that commonly inhabits the large intestines of warm blooded
animals. The host species provides Salmonella with a warm environment and supplies the
bacteria with nutrients such as amino acids and sugars (Davies and Wray, 1996). Salmonella
can get into the soil when manure and fertiliser is spread onto the soil. Salmonella is an
enteric or faecal organism that is found in the intestines of animals such as birds and
mammals. When these animals defecate the Salmonella is passed out of the body of the
animal and onto the soil. This may also cause the soil to become contaminated with
Salmonella. When the Salmonella is secreted from the host it battles with various
environmental conditions to survive. Salmonella in the soil can then be taken up into the
roots, or stems of fruits such as berries or vegetables such as carrots, potatoes and lettuce. To
survive in the soil Salmonella needs water, food source and the right conditions such as pH
25
and temperature. If manure is treated effectively then the Salmonella contamination can be
reduced so it is less applied to the soils. Manure is commonly treated by changing the pH and
temperature (Davies and Wray, 1996).
On the soil, Salmonella counts start to decrease and will eventually become low or non-
detectable. Salmonella has been found to survive in the soil for up to 900 days. Some
research has found that Salmonella will reach low levels over 30 days. It is recommended to
not use produce, roots or tubes between 30 days of the manure being spread on the soil. Birds
and insects such as flies are also very important vectors for the widespread transmission of
Salmonella in the environments (Davies and Wray, 1996).
The infection of wild bird populations with Salmonella has been linked with proximity to
farms and the incidence of human salmonellosis in the same area (Monaghan et al, 1985).
The common house fly can also be a carrier of Salmonella. This is a problem on dairy farms
and poultry farms. Salmonella survives in flies for up to 4 weeks. Flies inoculated with
Salmonella show a total defecation output of as much as 107 cfu. This is a problem as flies
that come into contact with manure food and water can transmit Salmonella.
Rapid transmission of Salmonella between the host species can also occur by the passage of
the bacteria from infected animals to fruit and vegetables due to the field being fertilised with
raw untreated manure. The detection of Salmonella on vegetables is linked with warm, moist
weather conditions during summer seasons. When Salmonella come into contact with
flowers, stems or fruits it colonizes the plant tissues.
Salmonella can be disseminated in soil even when fertiliser or manure is absent. This is a
result of water currents, rain runoff and underground springs which carry the contaminated
material onto the soil (Chao et al, 1987).
Salmonella spp. can survive in soils for 968 days. Survival times for up to 300 days in soils
spread with cattle slurry have been observed, with survival of 259 days reported for soils
amended with animal faeces (Jones, 1986). From a study conducted by Marsh et al (1998), S.
Typhimurium LT2 was detected from soil at a minimum concentration on 103 cells/ g.
Water sources:
Salmonella is frequently isolated from water sources. This source can aid the transmission
between hosts (Cherry et al, 1972). It is found that Salmonella can survive up to 15 days in a
26
septic tank system. Seepage can occur for the septic tanks onto the fields. Sewage and storm
runoff can also facilitate the bacterial passages into surface waters. Salmonella has a high
survival rate in aquatic environments. Wastewater and municipal wastes should only be used
if effective disinfecting systems are available. Surface water from streams and lakes can
contain pathogenic bacteria, protozoa and viruses which may lead to the contamination of
fruit and vegetable plants.
Manure:
The main causes of contamination includes animal manure or sewage waste. Manure should
be composted aerobically to reach 60-80 °C for a minimum of 15 days. Manure is often
applied to fields as fertiliser for fruit and vegetable production. Manure from pigs (or cattle)
is most commonly used. The application of untreated manure onto fields has a high risk of
contaminating the soil in the fields with harmful microorganisms such as E.coli and
Salmonella. According to a study conducted by Tauxe (1997) there is an increased
association between food-borne disease and fresh vegetable consumption in recent years.
Investigations have been conducted in some foodborne outbreaks linked to the consumption
of raw fruits and vegetables, and have identified manure as the primary source of
contamination.
Harvesting:
In an investigation of several foodborne illnesses associated with fresh produce (NACMCF,
1999) agricultural workers were in many cases the likely source of the pathogen.
Washing:
Washing of fruits vegetables during harvesting can remove soil and dirt. However, washing
can also be a source of microbial contamination. Chlorine can be used to disinfect and
prevent microorganisms growing during the sprouting of seeds (Sivapalasingam et al, 2000).
There are a wide range of agents available to disinfecting or sanitising fresh produce but their
efficiency can be variable and none of the disinfecting agents are able to ensure the
elimination of pathogens.
Retail:
27
A survey of fresh fruit and vegetables was conducted by Abadias et al (2008) in retail
establishments in Catalonia in Spain in 2006 to determine if the produce was contaminated
with pathogenic microorganisms. A survey of fresh and minimally-processed fruit and
vegetables, and sprouts was conducted in several retail establishments in the Lleida area
(Catalonia, Spain) during 2005–2006 to determine whether microbial contamination for the
fresh-cut vegetables that we analysed showed that, in general, the highest microorganism
counts were associated with grated carrot, arugula and spinach 7.8, 7.5 and 7.4 log cfu g−1 of
aerobic mesophilic microorganisms; 6.1, 5.8 and 5.2 log cfu g−1 of yeast and moulds; 5.9,
4.0 and 5.1 log cfu g−1 lactic acid bacteria and 6.2, 5.3 and 6.0 log cfu g−1 of
Enterobacteriaceae. The processing stages of fresh-cut products, such as handling, cutting,
shredding, slicing and grating, are all potential sources of contamination, which could further
increase the microbial load.
Handling and personal hygiene in retail environment can also cause contamination of fruits
and vegetables. Workers should use hairnets and clean outfits when handling the product.
The main source of contamination in terms of product preparation for the market is probably
water. Water is essential for packinghouse operations either for washing the product and
containers.
Packaging and transport:
Foreign materials may be found on products or inside the packaging of products (López
Camelo, 2002). These materials include dust, animal faeces, grease, oils, human hairs,
insects, and plant debris. This is usually due to insufficient care in handling. Packaging
environment can create favourable conditions for pathogens to live in. Other considerations
to prevent cross contamination include not storing or transporting fruits and vegetables with
other fresh food items. The atmosphere at which a product is stored also influences microbial
development.
Processing of ready to eat fruit and vegetables:
28
Cutting, slicing, skinning and shredding removes or damages the protective surfaces of the
plant or fruit (Garg et al, 1990). Cutters and slicers can be sources of contamination. They
can since provide inaccessible sites, which harbour bacteria. It may be very difficult to clean
these sites on the slicers and cutters so cross contamination of bacteria from the equipment
onto the fruit and vegetables can occur. The presence of cut surfaces on fruit and vegetables
can allow bacteria to enter into the produce. This is very hard to eliminate during washing.
Bruises and cuts allow the microbial infiltration of the tissues. Exposing vegetables to various
types of cutting has been shown to result in a six to seven-fold increase in microbial numbers.
29
6. Risk characterisation
Table 2: Summarised qualitative risk assessment for Salmonella spp. in ready-to-eat fruit and vegetables
FARM
Output required Summarised information Assessed probability of concentration and
key uncertainties
P6: The probability that the soil is contaminated
with Salmonella
C6: The concentration of Salmonella in the soil
P10: The probability that the fruit and vegetables
with be exposed to Salmonella during packaging
on the farm
C10: The concentration of Salmonella being
contracted during packaging
Pathogens can contaminate fruit and
vegetables during different points
throughout pre harvesting and post
harvesting systems
Potential pre-harvest sources of
microorganisms include soil, faeces,
irrigation water, dust, insects, manure,
wild and domestic animals, and human
handling
Infected animals are likely to excrete
Salmonella into the soil, into manure, or
onto the plant
The probability of Salmonella in soil is
High probability of seeds and soil being
contaminated.
If seeds and soil are contaminated
probability of fruit and vegetable plants
to be contaminated is high
If contamination occurs during washing
of packaging of produce the
concentration of Salmonella is likely to
be low
Key Uncertainties:
Frequency of faecal contamination
Frequency of human contamination to
fresh produce
Numbers of organisms on the fresh
30
high. If contamination does occur the
concentration of Salmonella found in the
soil is likely to be high. Salmonella is
found to survive for 900 days in the soil.
Washing, storage and packaging of
produce on the farm can cause
contamination
If contamination is to occur during
washing, this risk may be low
If contamination is to occur during
packaging the risk of contamination may
be low.
produce if contamination occurs
DISTRIBUTION & STORAGE
31
P12: The probability of the fruit and
vegetables becoming exposed to Salmonella
in the wholesale market
C12: The concentration of Salmonella present
in the wholesale market environment
P13: The probability that the fruit and
vegetables will contract Salmonella during
processing, such as cutting, slicing and dicing
C13: The concentration of Salmonella that
could be contracted by the produce in the
processing station
It is unlikely that Salmonella will
contaminate the fresh produce during
transportation and storage of the
produce to wholesale markets.
If the transport containers are cleaned
properly and good hygienic practices
are in place with handlers of the fruit
and veg then the risk of contamination
will be low.
If contamination does occur the
concentration of organisms is likely to
be low
Fresh produce can become
contaminated during the processing of
the produce. Processing involves
preparing the fruit and veg for sale as
ready-to-eat products
Contamination can occur during
processing due to contaminated
equipment or surfaces used for slicing,
dicing and washing of the produce.
Contamination during processing
There are 2 scenarios which could result in
contamination:
Scenario 1:
Fresh produce may be contaminated during
harvesting in the farm. If not washed
adequately the produce may contain high
numbers of organisms. These organisms are
the transported to wholesale markets and pose
a high risk of salmonellosis.
Scenario 2:
The produce may be properly disinfected
during farming and may become
contaminated during the processing stages,
due to unsanitary processing equipment.
If the produce is contaminated, the
concentration may be high. There is a high
risk of causing salmonellosis if produce is
contaminated during processing as the ready-
to-eat products are consumed usually without
32
stages is likely to be due to cross-
contamination due to poor hygienic
practices in place in the processing
plant. Cutters and slicers can be
sources of contamination. They can
provide inaccessible sites, which
harbour bacteria. It may be very
difficult to clean these sites on the
slicers and cutters so cross
contamination of bacteria from the
equipment onto the fruit and
vegetables can occur
If processing equipment is improperly
sanitised then there is a high risk of
contamination of produce during
processing. It is uncertain what the
concentration of organisms on the
fresh produce will be if contaminated
during processing.
any preparation.
Key uncertainties:
Frequency of post-harvesting
contamination
Frequency of contamination in
wholesale markets
Frequency of contamination during
processing due to contaminated
equipment.
RETAIL & HOME
33
P18: The probability of the fruit and
vegetables being exposed to Salmonella
during storage in retail facilities
C18: The concentration of Salmonella present
in storage containers in retail facilities
P20: The probability of the produce
becoming contaminated with Salmonella in
the consumer’s home.
C20: The concentration of Salmonella
contracted by the produce whilst present in
the consumers home
P: Probability of exposure of Salmonella per
serving
C: Number of organisms consumed per
serving
Fresh fruit and vegetables are unlikely
to be contaminated with Salmonella in
retail environments. If they are
contaminated with Salmonella then the
number of organisms present is likely
to be low.
The contamination of the fresh
produce is likely to occur from the
processing stages.
Possible sources of contamination in
the retail facilities can be due to
unclean storage containers of cross-
contamination by human handling of
produce
Fresh produce is unlikely to become
contaminated in the consumer’s home
if stored under correct conditions.
Uncertainty associated with
temperature and storage abuse in the
home
Uncertainty associated with hygiene
Probably a Low probability of
exposure per serving, this risk could
be higher if post harvesting and
processing contamination occurs.
Key uncertainties:
Storage conditions in retail or home
environment
Sources of contamination in
consumers home
Preparation of produce before
consumption
34
practices in home.
The organism is unlikely to grow if
produce is stored in clean containers
and under the correct conditions in
retail and home environments.
Probability of contamination in these
environments is likely to be low, but
could be high if hygiene practices are
not in place.
If they are contaminated in both
environments the concentration of
organisms could be low, but may be
high if contamination occurred pre and
post processing stages.
35
7. Discussion
Recommendations
Seed/ plant imports: To ensure that there is no risk of contamination at this stage , it would
be wise for importers/farmers who will be growing them to know where the products are
coming from and the hygiene/ disinfection procedures in place. It would also be ideal to have
a quarantine area for imported goods to avoid cross contamination with equipment and other
plants in the facility.
Seed/ plant transport: To avoid possible contamination, any imported seeds/plants should
be transported in sealable, easily sterilised containers. These could then be easily re-sterilised
if contamination was later found in the seeds/plants and the containers can be disinfected to
remove the possibility of affecting any produce thereafter.
Soil: To reduce the level of contamination in soils prior to growing of fruit and vegetables, it
is advised to treat soils, by influencing the temperature or pH.
s
Manure/irrigation water/ fertilisers: The Food Safety Authority has produced draft
guidelines for growers to minimise the risks of microbial contamination of RTE crops (FSA
2009). The guidelines point at a range of measures that can help kill pathogens that are
present in manures and slurries including:
exposure to sunlight and ultra violet rays
high temperatures (above 55°C)
low acid or high alkaline conditions (use of quick lime or slaked lime to raise pH
levels)
drying
the passage of time (though some bacteria such as E. coli can survive for several
months)
The draft guidance recommends a package of measures before, during and after the growing
season including:
careful selection of sites of fields
lay off periods between application of manures and slurries before harvesting
36
not allowing livestock to roam on lands where crops will soon be grown or harvested
recommendations for storing manures and slurries
the use of potable water for washing procedures
Water for irrigation: To avoid contamination from the water used to irrigate crops, it is
recommended that the water source be isolated off and covered to protect against becoming
contaminated from nearby livestock, run off from the land, heavy rainfall causing seeping of
contaminated water into the water source. Elevating the source above ground level will aid
with this immensely. Water should also be tested for contamination at regular, pre-specified
intervals.
Animals (including insects and avian species): It is recommended to keep livestock away
from areas where crops are to be grown and harvested by putting up physical barriers around
fields and crops as well as careful selection of sites for crops (also taking into consideration
the factor of runoff of faecal matter into the soil from heavy rainfall).
Harvesting of fruit/vegetables: There are many recommended steps that need be taken at
this stage in order to avoid contamination of the crops.
Direct contamination from field workers hands (if being manually harvested from fields) or
handling when being prepared for processing is a significant aspect of this process. So a
effective hygiene regime for workers is key to reduce this risk.
To prevent cross contamination during harvesting, thorough cleaning and decontamination of
equipment, containers and transport vehicles must be undertaken.
Washing of produce on farm (Food Safety Authority of Ireland, 2000) : Washing produce
reduces the chances of micro-organisms and chemicals remaining on the surface. This is an
essential step since most contaminants are on the surface of fruit and vegetables and can also
contaminate surrounding produce, thus spreading the hazard. Most post-harvest processes
involve a considerable degree of water to surface contact.
The following points should be considered:
The washing process must be sufficient to remove soil, chemicals, micro-organisms
and foreign bodies.
37
If the produce is not subject to bruising, then vigorous washing increases the chances
of removing the microbiological or chemical hazard. Surface scrubbing using brushes
is even more effective but only if the brushes are regularly cleaned and sanitised.
A series of washes is more effective than a single wash if the produce is washed with
fresh clean water each time. This prevents the build-up of micro-organisms and
washed-off chemicals in the wash water.
Wash water should be maintained below 10°C to reduce the growth rate of
microorganisms that are washed off the fresh produce. This is important if water is
not replaced frequently with clean water.
Wash water must be renewed frequently to prevent a build-up of micro-organisms and
preferably be treated with an effective antibacterial water treatment, such as chlorine,
with frequent monitoring to ensure effective concentrations are maintained.
The inclusion of a decontamination step to destroy micro-organisms should be
considered where appropriate. It is especially important where the quality of water
cannot be guaranteed or when produce items are intended to be eaten without further
cooking. However, a decontamination step may only be effective if produce is pre-
washed in potable water to remove as much soil as possible before decontamination.
This ensures that the decontamination step, using water treated with an antibacterial
treatment like chlorine, remains effective. Organic matter inactivates chlorine.
Storage of produce on farm: To avoid contamination on the farm when storing produce, it
is essential that good hygiene practices are upheld so that there are no pathogenic bacteria
(for example, Salmonella) left on the surfaces of the fruit and vegetables and the containers
that they are being stored in so that they can grow and proliferate during storage at ambient or
refrigeration temperatures, both at which Salmonella can grow, abet sub-optimally.
Packaging on farm: (Safefoods, 2007) Modified Atmospheric packaging (MAP) is defined
as an atmosphere created by altering the normal composition of air to provide an atmosphere
capable of extending shelf-life. In MAP, gases such as oxygen, carbon dioxide and nitrogen
are used to alter the composition of the atmosphere around the product so that the storage life
can be extended. The product is then sealed in a wrap like polyethylene, polypropylene,
polyvinyl chlorine or edible film. This is the optimal packaging type for storing fresh
produce in as it reduces risk of pathogenic bacteria proliferating on the crops.
38
Tissue disruption caused by processing results in elevated respiration and transpiration, which
can lead to rapid deterioration. In addition, cut tissues release nutrients that support the
growth of micro flora present on raw produce. The O2 level in packs of fruit or vegetables is
usually kept between one and five per cent, which will reduce the respiration rate and,
therefore, oxidative breakdown of fruits and vegetables (Lee, Arul et al. 1995). Respiration
uses O2 and typically produces CO2 therefore making packages anaerobic. O2 levels below
eight per cent also reduce the level of ethylene, which delays ripening and maturation.
However, low levels of O2 can increase anaerobic respiration and sensory degradation.
Given that MAP alone is not sufficient to prevent pathogen growth, chilling is extremely
important and Hazard Analysis Critical Control Point (HACCP), Good Manufacturing
Practice (GMP) and Good Agricultural Practice (GAP) should be in place to prevent
pathogen contamination.
Transport of fruit and vegetables: As recommended before with seeds/plants, packaged
goods should be transported in sealable, easily sterilised containers. This helps prevent
possible contamination from the environment and from the transport vehicle itself. It also
protects the products when being moved around and transferred to warehouses and retail
locations. These could then be easily re-sterilised and can be disinfected to remove the
possibility of affecting any produce thereafter. Fresh fruit and vegetables also need to be
transported in chilled vehicles so as to preserve them and reduce risk of contaminates
growing.
Wholesale market: Even though the produce is usually packaged and sealed at this point, it
is still crucial for wholesalers to be as vigilant as if the produce was uncovered. If handled
improperly, fruits and vegetables can be crushed or damaged, allowing for easy proliferation
of pathogenic bacteria. The transport vehicles that are used to distribute the produce must be
cleaned down and decontaminated as standard and be refrigerated to reduce bacterial growth.
Processing station:
Preliminary washing: To avoid contamination, produce that comes into a processing plant
needs to be washed to remove any remaining debris, soil and possible contaminants on the
surface of the fruit or vegetables. Also, potable or disinfected water must be used so as not to
add any more bacteria onto the surfaces of the produce.
39
Slicing or shredding: Once any stalks and outer leaves are trimmed and removed from
vegetables, they are sliced or shredded, usually using machinery for both steps. Given this, all
machinery must be disinfected and cleaned regularly and very thoroughly in order to avoid
microbial build up, growth and subsequent cross contamination into the internal tissues of the
fruits and vegetables, which are optimal for bacteria growth and proliferation.
Washing/Disinfection: Again, as mentioned previously, washing is a key step to remove
possible contaminants on the surface of the produce. Bacteria can also be internalised via
waterborne contamination. This can occur when fruits are put into a wash tank and water is
taken up into fruits, particularly when the fruits are warm and the wash water is cold.
Therefore, clean, potable water must be used.
Internalisation of contaminants into the internal tissues is a significant problem as washing
will not remove these contaminants. Great care needs to be taken at this step, along with all
previous and subsequent steps to avoid this.
An effective decontamination stage is essential prior to packaging to help reduce the level of
pathogenic and spoilage organisms in RTE produce. The most common substance used for
decontamination is chlorine, however, care must be taken so that appropriate concentrations
are used and that organic matter is completely removed from the produce as it would deem
the chlorine ineffective.
Packaging: As with packaging on the farm, the same procedures can be used for processing
for retail. A comprehensive strategy should be employed at this stage so as to keep
uncontaminated produce free from contamination or produce that has contaminated can be
preserved so as to avoid growth and proliferation. Use of packaging such as MAP along with
storage at refrigerated temperatures is therefore essential here.
Retail storage: Fresh produce needs to be stored at refrigerated temperatures of 2-5°C.
Ideally, they must be kept separately, or stored at the top of the fridges if being stored along
with other raw foods, as they can cause cross contamination if being stored improperly. This
is key especially since a large portion of fruits and salad vegetables are not subjected to
another heating step, so pathogenic contaminants cannot be killed at this stage in the
pathway.
Consumer: In the case of ready-to-eat (RTE) fruits and vegetables, they are often sold in
individual portions so make them perfect for eating straight away, marketed as a on the go
40
snack or lunch accompaniment. This being said, many people do not wash or treat them in
any way, so it is a notable source of disease from pathogen bacteria present in RTE products,
such as Salmonella.
To avoid this, consumers should be advised to wash fruits and vegetables, even if pre-
packaged.
Storage at home: In a domestic setting, this is a crucial step to not only prevent
contamination of RTE fruit and vegetables but can be where contamination can happen
frequently if they are stored improperly. To avoid contamination, these products need to be
stored, refrigerated, separately from raw produce such as raw meats, either in a chilled drawer
or in sealed containers so that any liquids cannot drop into them or that they do not touch and
become cross contaminated. Good hygiene of the fridge is also essential, since there are
pathogenic bacteria that can persist and survive at refrigeration temperatures and begin to
grow once they come in contact with foods.
Preparation before use: To avoid contamination, fresh fruit and vegetables that will not
undergo any further cooking must be prepared separately to raw meats, especially chicken in
regard to Salmonella contamination. Hygiene of the person preparing the foods as well as all
the utensils used is of the greatest importance, as it is through those that contamination occurs
and leads to food-borne gastroenteritis. Knowledge of effective procedures for hand washing
as well as decontamination of tools, utensils and containers must be in place, even at a
domestic level.
8. Conclusions
41
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