Microbes and Infection 4 (2002) 413–423

www.elsevier.com/locate/micinf

Current focus

Ecological factors influencing survival and growth of human pathogens

on raw fruits and vegetables
Larry R. Beuchat *
Center for Food Safety and Department of Food Science and Technology, University of Georgia, 1109 Experiment Street, Griffın, GA 30223-1797, USA

Abstract

Outbreaks of human infections associated with consumption of raw fruits and vegetables have occurred with increased frequency during
the past decade. Factors contributing to this increase may include changes in agronomic and processing practices, an increase in per capita
consumption of raw or minimally processed fruits and vegetables, increased international trade and distribution, and an increase in the
number of immuno-compromised consumers. A general lack of efficacy of sanitizers in removing or killing pathogens on raw fruits and
vegetables has been attributed, in part, to their inaccessibility to locations within structures and tissues that may harbor pathogens.
Understanding the ecology of pathogens and naturally occurring microorganisms is essential before interventions for elimination or control
of growth can be devised. © 2002 Éditions scientifiques et médicales Elsevier SAS. All rights reserved.

Keywords: Fruits; Vegetables; Human pathogens

1. Introduction bials that may enhance or retard the growth of naturally

occurring microflora and pathogens [6]. The presence of soil
The number of documented outbreaks of human infec- or fecal material on the surface of produce that may
tions associated with consumption of raw fruits, vegetables, permeate cut tissues may alter the ecological environment
and unpasteurized fruit juices has increased in recent years and, perhaps, also the behavior of pathogens and other
[1–3]. Advances in epidemiologic surveillance programs microflora. The growth of molds in these environments may
have enabled these associations to be made. However, result in increased pH, thus enhancing the probability of
changes in dietary habits, methods of fruit and vegetable growth of pathogenic bacteria. Colonization and biofilm
production and processing, sources of produce, and the development may ensue, resulting in conditions that would
emergence of pathogens previously not recognized for their protect against death of pathogens or promote growth of
association with raw produce have enhanced the potential spoilage or pathogenic microorganisms. The viability of
for outbreaks [4,5]. parasites as affected by extrinsic and intrinsic factors unique
While much is known about the ecology of microbial to fruits and vegetables is unknown.
pathogens in foods of animal origin, the behavior of
pathogens in association with naturally occurring microflora
on fruits and vegetables is ill-defined. Tremendous differ-
ences in surface morphology, internal tissue composition, 2. Pathogens isolated from or associated with fruits
and metabolic activities of leaves, stems, florets, fruits, and vegetables
roots, and tubers provide a wide range of diverse ecological
niches selective for specific species or groups of microor- Although spoilage bacteria, yeasts, and molds dominate
ganisms. the microflora on raw fruits and vegetables, the occasional
Bruised and cut surface tissues exude fluids containing presence of pathogenic bacteria, parasites, and viruses
nutrients and numerous phytoalexins and other antimicro- capable of causing human infections has also been docu-
mented (for reviews see [2,7–12]). All types of produce
* Corresponding author. Tel.: +1-770-412-4740; fax: +1-770-229-3216. have potential to harbor pathogens [13] but Shigella spp.,
E-mail address: [email protected] (L.R. Beuchat). Salmonella, enterotoxigenic and enterohemorrhagic Es
© 2002 Éditions scientifiques et médicales Elsevier SAS. All rights reserved.
PII: S 1 2 8 6 - 4 5 7 9 ( 0 2 ) 0 1 5 5 5 - 1
414 L.R. Beuchat / Microbes and Infection 4 (2002) 413–423

cherichia coli, Campylobacter spp., Listeria monocytoge- or greenhouses, or during harvesting, post-harvest handling,
nes, Yersinia enterocolitica, Bacillus cereus, Clostridium processing, distribution, and preparation in food service or
botulinum, viruses, and parasites such as Giardia lamblia, home settings. Listed in Table 1 are examples of fresh
Cyclospora cayetanensis, and Cryptosporidium parvum are vegetables from which pathogenic bacteria have been iso-
of greatest public health concern [2,7,14,15]. Fruits and lated. Each vegetable in this list possesses a unique set of
vegetables can become contaminated with pathogenic mi- intrinsic factors that can influence the survival and growth
croorganisms while growing in fields, orchards, vineyards, of human pathogenic microorganisms.
Table 1
a
Examples of pathogenic bacteria isolated from raw vegetables
Vegetable Country Pathogen Prevalence b Reference
Alfalfa sprouts USA Aeromonas [16]
USA S. Meleagridis [17]
USA Bacillus cereus [18]
Alfalfa seeds USA S. Havana [19]
S. Cubana
S. Tennessee
USA S. Newport [20]
Denmark
Artichoke Spain Salmonella 3/25 (12%) [21]
Asparagus USA Aeromonas [22]
Bean sprouts Malaysia L. monocytogenes 6/7 (85%) [23]
Sweden Salmonella [24]
Thailand Salmonella 30/344 (8.7%) [25]
Beet leaves Spain Salmonella 4/52 (7.7%) [21]
Broccoli USA Aeromonas [26]
USA Aeromonas 5/16 (31.3%) [16]
Cabbage Canada L. monocytogenes 2/92 (2.2%) [27]
Mexico E. coli O157:H7 1/4 (25.0%) [28]
Peru Vibrio cholerae [29]
Spain Salmonella 7/41 (17.1%) [21]
Sri Lanka L. monocytogenes 6/18 (33%) [30]
USA L. monocytogenes 1/92 (1.1%) [31]
Carrots Lebanon Staphylococcus (14.3%) [32]
Cauliflower Netherlands Salmonella 1/13 (7.7%) [33]
Salmonella 1/23 (4.5%) [21]
Celery USA Aeromonas [26]
Mexico E. coli O157:H7 6/34 (17.6%) [28]
Chili Spain Salmonella 2/26 (7.7%) [21]
Cilantro Surinam Salmonella 5/16 (31.3%) [33]
Coriander Mexico E. coli O157:H7 8/41 (19.5%) [28]
Cress sprouts Mexico E. coli O157:H7 2/20 (20.0%) [28]
Cucumber USA B. cereus [18]
Malaysia L. monocytogenes 4/5 (80%) [23]
Pakistan L. monocytogenes 1/5 (6.7%) [34]
Egg plant USA L. monocytogenes 2/92 (2.2%) [31]
Endive Netherlands Salmonella 2/13 (1.5%) [33]
Fennel Netherlands Salmonella 2/26 (7.7%) [33]
Green onion Italy Salmonella 4/89 (71.9%) [35]
Leafy vegetables Canada Campylobacter 1/40 (2.5%) [36]
Lettuce Malaysia L. monocytogenes 5/22 (22.7%) [23]
Canada Campylobacter 2/67 (3.1%) [36]
Italy Salmonella 82/120 (68.3%) [35]
Lebanon Staphylococcus (14.3%) [32]
Malaysia L. monocytogenes 1/28 (3.6%) [37]
Netherlands Salmonella 2/28 (7.1%) [33]
Spain Salmonella 5/80 (6.3%) [21]
Sri Lanka L. monocytogenes 10/20 (50%) [30]
USA Aeromonas [16]
Mungbean sprouts UK S. Saint-Paul [38]
Mushrooms US C. jejuni 3/200 (1.5%) [39]
Mustard cress UK S. Gold-Coast [40]
Mustard sprouts USA B. cereus [18]
Parsley Canada Campylobacter 1/42 (2.4%) [36]
Egypt Shigella 1/250 (0.4%) [41]
Lebanon Staphylococcus (7.7%) [32]
Spain Salmonella 1/23 (4.3%) [21]
 L.R. Beuchat / Microbes and Infection 4 (2002) 413–423 415

Vegetable Country Pathogen Prevalence b Reference

Pepper Canada Campylobacter 1/63 (1.6%) [36]
USA Aeromonas [16]
Sweden Salmonella [42]
Potatoes USA L. monoyctogenes 19/70 (27.1%) [31]
USA L. monocytogenes 28/132 (21.2%) [43]
Prepacked salads Canada Campylobacter 2/74 (2.7%) [36]
Northern Ireland L. monocytogenes 3/21 (14.3%) [44]
UK L. monocytogenes 4/60 (13.3%) [45]
Radish Lebanon Staphylococcus (6.3%) [32]
USA L. monocytogenes 25/68 (36.8%) [31]
USA L. monocytogenes 19/132 (14.4%) [43]
Salad greens Egypt Salmonella 1/250 (0.4%) [41]
UK S. aureus 13/256 (5.1%) [46]
Salad vegetables Egypt Shigella 3/250 (1.2%) [41]
Egypt S. aureus 3/36 (8.3%) [32]
Germany L. monocytogenes 6/263 (2.3%) [47]
Northern Ireland L. monocytogenes 4/16 (25%) [44]
UK Y. enterocolitica 4/16 (25%) [48]
UK L. monocytogenes 2/108 (1.8%) [49]
Seed sprouts Canada Staphylococcus 13/45 (24%) [50]
Soybean sprouts USA B. cereus [18]
Spinach Canada Campylobacter 2/60 (3.3%) [36]
Spain Salmonella 2/38 (5.2%) [21]
USA Aeromonas [16]
Sprouting seeds USA B. cereus 56/98 (57%) [51]
Tomato Pakistan L. monocytogenes 2/15 (13.3%) [34]
Vegetables Egypt Salmonella 2/250 (0.8%) [41]
France Y. enterocolitica 4/58 (7%) [52]
France Y. enterocolitica 15/30 (50%) [53]
Iraq Salmonella 3/43 (7.0%) [54]
Italy L. monocytogenes 7/102 (6.9%)
Italy Y. enterocolitica 1/102 (1.0%) [55]
Spain L. monocytogenes 8/103 (7.8%) [56]
Spain Salmonella 46/849 (5.4%) [57]
Taiwan L. monocytogenes 6/49 (12.2%) [58]
UK L. monocytogenes 4/64 (6.2%) [59]
USA Salmonella 4/50 (8.0%) [60]
.
a
Adapted from [7] and used with permission of the International Association of Food Protection.
b
Number of samples positive out of number analyzed; percentage of positive samples is in parenthesis.

A wide range of fresh fruits and vegetables, as well as scale have enabled the raw fruit and vegetable industry to
unpasteurized fruit juices, has been implicated in outbreaks supply consumers with a wide range of high-quality pro-
of infections. Examples are listed in Table 2. This is not a duce in most countries year round. Some of the same
comprehensive list but does illustrate the diversity of types technologies and practices have also introduced an in-
of produce potentially capable of serving as vehicles for creased risk for human illness associated with pathogenic
human infection. The survival and growth of a pathogen on microorganisms. The use of animal manure that has not
or in raw produce or unpasteurized produce products are been composted rather than chemical fertilizer, as well as
dictated by its metabolic capabilities. However, manifesta- untreated sewage or irrigation water containing pathogens
tion of these capabilities can be greatly influenced by contributes to this risk. Changes in social demographics,
intrinsic and extrinsic ecological factors naturally present in food consumption patterns, and awareness by epidemiolo-
produce or imposed at one or more points during the entire gists and health care professionals that raw fruits and
system of production, processing, distribution, and prepara- vegetables are potential vehicles of infections may also be
tion at the site of consumption. contributing to an increase in documented produce-
associated outbreaks of human illness.
Many of the factors that are thought to contribute to the
epidemiology of diseases associated with raw fruits and
3. Possible reasons for increased numbers of outbreaks vegetables [5,110] are directly or indirectly impacted by
or infections ecological conditions that affect survival or growth of
pathogenic microorganisms. Adaptation to stress environ-
Changes in agronomic, processing, preservation, packag- ments can result in a pathogen becoming better suited to
ing, distribution, and marketing technologies on a global survival and growth, or to becoming more virulent. E. coli
416 L.R. Beuchat / Microbes and Infection 4 (2002) 413–423

Table 2
Examples of outbreaks of infections epidemiologically associated with raw fruits and vegetables and unpasteurized products
Microorganism Year Location Type of produce or product Reference
Bacteria
Bacillus cereus 1973 USA Seed sprouts [61]
Clostridium botulinum 1987 USA Cabbage [62]
E. coli O157:H7 1991 USA Apple cider [63]
1995 USA Lettuce [64,65]
1996 USA Apple juice [66,67]
1997 Japan Radish sprouts [68]
1997 USA Alfalfa sprouts [69]
E. coli (enterotoxigenic) 1993 USA Carrots [70]
Listeria monocytogenes 1979 USA Celery, lettuce, tomato [71]
1981 Canada Cabbage [72]
Salmonella
Miami 1954 USA Watermelon [73]
Typhimurium 1974 USA Apple cider [3]
Oranienburg 1979 USA Watermelon [74]
Saint-Paul 1988 UK Mungbean sprouts [75]
Chester 1989-90 USA Cantaloupes [76]
Javiana 1990 USA Tomatoes [77]
Poona 1991 USA/Canada Cantaloupes [78]
Montevideo 1993 USA Tomatoes [5]
Bovismorbificans 1994 Sweden/Finland Alfalfa sprouts [79]
Hartford/Gaminara/Rubislaw 1995 USA Orange juice [80]
Stanley 1995 USA Alfalfa sprouts [81]
Montevideo/Meleagridis 1996 USA Alfalfa sprouts [17]
Typhi 1998-99 USA Mamey [82]
Mbandaka 1999 USA Alfalfa sprouts [83]
Shigella flexneri 1998 UK Fruit salad [84]
S. sonnei 1986 USA Lettuce [85]
1994 Norway Lettuce [86]
1998 USA Parsley [87]
1995 USA Scallions [88]
Vibrio cholerae 1970 Israel Vegetables [89]
1991 USA Coconut milk [90]
Viruses
Calicivirus 1998 Finland Raspberries (frozen) [91]
Hepatitis A 1983 UK Raspberries (frozen) [92]
1988 USA Lettuce [93]
1990 USA Strawberries (frozen) [94]
1994 USA Tomatoes [95]
1997 USA Strawberries [96]
Norwalk and Norwalk-like 1987 UK Melon [97]
1982 USA Green salad [98]
1991 USA Celery [99]
Parasites
Cyclospora cayetanensis 1996-97 USA, Canada Raspberries [100–104]
1997 USA Lettuce [105]
1997 USA Basil/basil-containing products [106]
1997 Peru Raw vegetables [14]
Cryptosporidium parvum 1993 USA Apple cider [107]
1995 USA Mixed salad with celery [108]
1996 USA Apple cider [67]
1997 Peru Raw vegetables [14]
Giardia lamblia 1992 USA Raw vegetables [109]
.

O157:H7 [111–113] and Salmonella [114,115], for example, central processing facilities and subsequent storage and
are known to adapt to reduced pH and subsequently exhibit handling practices in food preparation areas may also be
increased tolerance to stress environments. Global trade and contributing to an increased frequency of produce-
international travel have resulted in increased contact of associated infections. The ecological behavior of pathogens
people with pathogens to which they had not been previ- and spoilage microorganisms on raw fruits and vegetables
ously exposed. Trends toward greater geographic distribu- can be greatly affected by these changes, thus resulting in
tion of minimally processed fruits and vegetables from increased risk of illness.
 L.R. Beuchat / Microbes and Infection 4 (2002) 413–423 417

4. Understanding the ecosystem of pathogens is that have been associated with the consumption of raw
paramount to devising methods for control vegetables are likely, in part, due to contamination by
manure from ruminants [8]. L. monocytogenes is known to
A better understanding of microbial ecosystems on the grow on a variety of vegetables at refrigeration temperatures
surface of raw fruits and vegetables would be extremely [13,125,126].
useful when developing interventions to minimize contami- Although produce may become contaminated with patho-
nation, prevent the growth of pathogens, and kill or remove gens as a result of contact with manure used as a soil
pathogens at various stages of production, processing, fertilizer, through manure-contaminated irrigation water, or
marketing, and preparation for consumption. These ecosys- by direct contact with feces from birds and grazing animals,
tems are extremely diverse and complex. The presence and very little is known regarding the effect of ecological
numbers of bacteria, yeasts, molds, parasites, and viruses conditions on survival of pathogens enmeshed in manure on
differ, depending on the type of produce, agronomic prac- the surface of fruits and vegetables. Likewise, although the
tices, geographical area of production, and weather condi- use of manure in the production of fruits and vegetables
tions prior to harvest [13,116,117]. Microbial ecosystems should be carefully managed to limit the potential for
unique to various types of produce after harvesting can be contamination with pathogens during pre-harvest, there are
greatly influenced by handling and storage conditions as few scientific data to validate practical treatments to kill
well as conditions of processing, packaging, distribution, pathogens in manure that may adhere to fresh produce.
and marketing. Intrinsic as well as extrinsic factors determine the range
Pathogens, along with spoilage microorganisms, may and populations of microorganisms associated with fruits
contaminate fruits and vegetables via several different and vegetables at any given point throughout their produc-
routes and at several points throughout the pre- and post- tion and post-harvest handling, thus influencing the rate and
harvest system. Potential pre-harvest sources of microor- type of spoilage that eventually renders raw produce ined-
ganisms include soil, feces, irrigation water, water used to ible. Surfaces of fruits, stems, roots, florets, and leaves, for
apply fungicides and insecticides, dust, insects, inad- example, are each characterized by unique microenviron-
equately composted manure, wild and domestic animals, ments that influence colonization of bacteria, yeasts, and
and human handling. Post-harvest sources include feces, molds, as well as attachment of these microorganisms,
human handling, harvesting equipment, transport contain- parasites, and viruses. The environment in which plants are
ers, wild and domestic animals, insects, dust, rinse water, grown imposes extrinsic factors that influence survival and
ice, transport vehicles, and processing equipment. Jan- growth of associated surface microflora, whereas intrinsic
isiewicz et al. [118] demonstrated that fruit flies contami- parameters such as the nature of the epithelium and protec-
nated with a fluorescent-tagged nonpathogenic strain of tive cuticle, tissue pH, and the presence of antimicrobials
E. coli O157:H7 served as a vector in colonization of the dictate which groups of produce may be more likely than
organism in apple wounds. The bacterium was isolated from others to harbor certain types of microorganisms in dam-
apple wounds within 48 h of exposure of apples to the flies. aged tissues.
Manure used as a fertilizer or soil amendment, as well as The range of microorganisms recovered from raw fruits
in irrigation water, represents a potential source of patho- and vegetables at harvest most often reflects the microflora
gens to contaminate fruits and vegetables. E. coli O157:H7 present in the field, orchard, grove, or vineyard at the time
and Salmonella are carried by animals and shed in their of harvest [2,117,127]. Climatic and agricultural determi-
feces [119,120]. Noncomposted or improperly composted nants affecting the microbial ecosystem at harvest include
manure used on the farm, or manure that enters surface geographical location, history of precipitation, wind, irriga-
waters, may contain these pathogens and subsequently tion practices, pre-harvest, harvest, and post-harvest prac-
contaminate produce [121]. Studies on the fate of E. coli tices, and the presence of insects, animals, and birds [4,13].
O157:H7 in bovine feces revealed that the pathogen sur- Gram-negative bacteria dominate the microflora associated
vived in cattle manure for 42–49 d at 37 °C, and for 49–56 d with most vegetables, while molds and weakly fermentative
at 22 °C [122]. Another study on the behavior of E. coli yeasts often comprise the majority of microflora on raw
O157:H7 in manure revealed that the pathogen could fruits, largely due to the acidic pH of fruit tissue, which
survive up to 47 d, 4 months, and 21 months, in bovine generally is less than 4.0 [128].
manure, aerated ovine manure, and nonaerated ovine ma-
nure, respectively [123]. Beuchat [124] was able to detect
E. coli O157:H7 on manure-contaminated lettuce stored at 5. Cohabitation with other microorganisms may affect
4 °C for up to 15 d, even when the initial inoculum was only survival and growth of pathogens
100-101 CFU/g.
L. monocytogenes is widely distributed in the environ- While the pH of many vegetables is in a range suitable
ment, where it is associated with decaying vegetation, soil, for growth of pathogenic bacteria, some, e.g. fully ripe
sewage, and feces of animals, and has been isolated from tomatoes, are in a pH range (3.9–4.4) that prevents or
several types of vegetables [7]. Cases of human listeriosis retards growth. Yeasts and molds, on the other hand, have a
418 L.R. Beuchat / Microbes and Infection 4 (2002) 413–423

competitive advantage over bacteria that may access bruised apples prevented E. coli O157:H7 from growing. Popula-
tissues of acidic vegetables and many fruits, because they tions of yeasts and molds on test produce were not reported
are able to grow at the lower pH range (2.2–5.0) character- in these studies [135,136].
istic of much of this produce. Spoilage of fruits is often With the exception of some types of melons, e.g. canta-
caused by specific molds or groups of molds and yeasts loupe (pH 6.2–6.9) and watermelon (pH 5.2–5.7), which are
[128]. When surface tissues of fruits are punctured or recognized as good substrates for growth of Salmonella
broken by insects or mechanical abuse, yeasts and molds [137] and E. coli O157:H7 [138], fruits and fruit juices with
naturally present on the skin surface can rapidly grow in the pH less than 4.0 are generally not considered as substrates
abundance of nutrients available in the released cell fluids. to support the growth of pathogenic bacteria. However, the
Spoilage by yeasts usually results from fermentative development of a pH gradient surrounding mycelial growth
activity. Molds, many of which can utilize ethanol and in bruised tissues or as a mat on the surface of juice could
simple sugars as sources of energy, then grow and eventu- provide conditions for growth of incident cells of patho-
ally degrade structural polysaccharides. Many molds pro- genic bacteria.
duce ammonia and other alkaline by-products during the Most of the natural microflora on the surface of fresh
course of metabolizing substrate nutrients. Some molds and produce do not exert a deleterious effect on sensory quali-
yeasts utilize organic acids, leading to reduced acidity and ties. However, when spoilage does occur, Pseudomonas,
increased pH. At least two reports show that growth of Xanthomonas, Erwinia, Bacillus, Clostridium, and several
molds on the surface of tomato juice (pH 4.2) increases the genera of yeasts and molds are commonly involved
pH to 6.3–7.6, allowing C. botulinum to grow and produce [139,140]. Some naturally occurring microorganisms may
toxin [129,130]. Of the 58 species representing 21 genera of have a lethal or antagonistic effect on bacteria capable of
molds examined by Mundt [131], all except two raised the causing human diseases. Bacteriocinogenic strains of Pedio-
pH of tomato juice (pH 4.1) to a range of 4.9–9.0. coccus and Enterococcus, for example, have recently been
Insects such as the lesser mealworm and house fly have shown to control the growth of L. monocytogenes on mung-
been shown to carry E. coli O157:H7 [132,133]. Fruits flies bean sprouts [141].
have been shown to transmit E. coli O157:H7 to apples
[118]. Recognizing that outbreaks of E. coli O157:H7 infec-
tions associated with apple cider may have been due, in part, 6. Behavior of pathogens in biofilms
to amplification of the pathogen within bruised apple tissue,
Dingman [134] investigated survival and growth of E. coli Means by which pathogens contaminate fresh produce
O157:H7 in tissue of five apple cultivars. While growth are several, including environmental sources in the field or
occurred in bruised tissue of all five cultivars, initiation of orchard, as noted above, or contact with harvesting equip-
growth varied from 2 to 6 d after inoculation and was ment and containers used to transport produce from the field
influenced by the time elapsed between picking and inocu- to the marketplace, and perhaps in food service and home
lating apples. Bacteria other than E. coli O157:H7 were not settings. Exopolysaccarides secreted by bacteria can form a
isolated from inoculated bruised tissue. Yeast and mold bound capsule layer when associated with the cell wall or
populations were not determined; however, the pH of released by the cell to create a matrix structure [142].
bruised apples was significantly higher than the pH of Microbial aggregates that may harbor bacteria, yeasts, and
undamaged apples, suggesting that mold growth may have molds within this matrix have been observed on plant
occurred. In any case, the increased pH would favor surfaces, and these structures are referred to as biofilms
survival and growth of E. coli O157:H7 and other patho- [143].
gens. Colonization of spoilage and non-spoilage microorgan-
In a survey of 401 samples of raw fruits and vegetables isms of fruits, vegetables, and post-harvest contact surfaces
collected in retail markets, 66% affected by bacterial soft rot can provide a protective environment for pathogens, reduc-
were positive for presumptive colonies of Salmonella [135]. ing the effectiveness of sanitizers and other inhibitory
Thirty percent of 166 representative isolates from 20 differ- agents [144]. L. monocytogenes, in a multispecies biofilm
ent commodities, including cantaloupe and tomato, were containing Pseudomonas fragi and Staphylococcus xylosus,
confirmed to be Salmonella. Co-inoculation of potato, has been reported to be essentially unaffected by treatment
carrot, and pepper with a soft-rot bacterium and with with 500 ppm free chlorine [145]. Fett [146] examined the
Salmonella typhimurium, followed by incubation for 24 h, cotyledons, hypocotyls, and roots of alfalfa, broccoli,
resulted in 10-fold higher counts of the pathogen compared cloves, and sunflower sprouts. Biofilms were observed on
to vegetables inoculated with Salmonella alone. Vegetables plant parts. He concluded that naturally occurring biofilms
co-inoculated with Pseudomonas viridiflava and S. typhimu- on sprouts may afford protected colonization sites for
rium contained Salmonella populations approximately three human pathogens such as Salmonella and E. coli O157:H7,
times higher than vegetables inoculated with Salmonella making their elimination with antimicrobial compounds
alone. Janisiewicz et al. [136], on the other hand, reported difficult. The formation of biofilms on leaf surfaces of
that inoculation of Pseudomonas syringae into wounds of spinach, lettuce, Chinese cabbage, celery, leek, basil, pars
 L.R. Beuchat / Microbes and Infection 4 (2002) 413–423 419

ley and endive has been demonstrated [143]. Estimates of O157:H7 has been described. Once positioned in the these
biofilm abundance in phyllosphere communities show that ecological niches, cells may survive and grow to high
bacteria in biofilms constitute 10–40% of the bacterial population by the time the infected produce is consumed.
population on broad-leaf endive and parsley [147]. Infiltration of pathogens into fruit and vegetable tissues is
Containers used to harvest, transport, and display raw dependent upon temperature, time, and pressure, and only
fruits and vegetables are often not effectively cleaned and occurs when the water pressure on the produce surface
sanitized, which can lead to the development of biofilms overcomes internal gas pressure and the hydrophobic nature
[148–150]. Even single-use containers may hold produce of the produce surface [158,159,163]. Infiltration is en-
for a sufficient time to allow the formation of biofilms. hanced if the temperature of the fruit or vegetable is higher
Contamination of fresh produce with pathogens may result than the temperature of a water suspension of cells. Addition
from contact with surfaces harboring these biofilms. If of detergents (surfactants) to water also promotes infiltra-
pathogens attach to biofilms during transport or processing, tion of produce, apparently by reducing the surface tension
their survival and growth may be enhanced [151–153]. of the water at the air–water interface with damaged cutin,
Growth of pathogens incorporated into biofilms would parenchemal cells, or pores leading into the tissues. Regard-
increase the probability of cross-contamination of produce. less of the mode of infiltration, cells may establish micro-
Jeong and Frank [154,155] determined that L. monocytoge- colonies that are extremely difficult to reach with aqueous
nes grows in multispecies biofilms containing microflora chemical solutions. This can result in a higher risk of
from meat and dairy plants. No information is available on consuming raw fruits and vegetables that may harbor
the behavior of L. monocytogenes or other pathogenic pathogenic microorganisms.
bacteria in biofilms formed by microflora associated with
raw fruits and vegetables. Predominant microorganisms in
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