Food poisoning outbreaks caused by fresh salad vegetables have been making food safety headlines on a regular basis in recent years, especially in the USA.
A series of large, high profile Salmonella and E. coli O157 outbreaks linked to spinach and lettuce has damaged the US fresh produce industry and left the sector with a reputation for high-risk products. But is that reputation really deserved and what can be done to prevent contamination and improve food safety in the salad sector? Research is beginning to reveal complex relationships between microbes and plants, which seem to play an important role in the contamination process. In the future, it may be possible to use this knowledge to devise innovative new ways to minimise, or even prevent, contamination with foodborne pathogens.
In September 2006 an outbreak of E. coli O157:H7 infection in the USA affected more than 200 people across 26 states. Nearly a third of those affected developed potentially serious complications, such as haemolytic uraemic syndrome (HUS), and at least one person died. Investigators soon discovered that the source of the outbreak was rather unusual. E. coli O157:H7 is usually associated with meat products, but case-control studies on this occasion pointed clearly to a single brand of fresh bagged baby spinach. Microbiological tests confirmed the investigators’ suspicions, as the outbreak strain was isolated from unopened bags of the implicated spinach. Consequently, consumers were advised not to eat the product and the producer recalled all the affected spinach.
The extent of the problem
The outbreak described above, plus many others in North America and elsewhere, have tarnished the safety image of prepared salads. This has coincided with a dramatic rise in demand for these products in response to healthy eating campaigns. The US annual market for prepared salads was estimated at $1.2 billion in 2005 before the spinach outbreak and a conservative estimate of the UK market came in at £256 million for the same year. Salads are big business, but are they also big contributors to foodborne disease?
A recent comprehensive review by the UK Health Protection Agency of prepared salad safety provides some of the answers. The review authors looked at more than 2000 general outbreaks of foodborne disease that were recorded in England and Wales from 1992 to 2006 and found that only 82 (4%) of them were associated with prepared salads. The review also pointed out that most of the outbreaks linked to salads occurred in the catering sector and were associated with infected food handlers, cross contamination and poor storage. Just two outbreaks were associated with salad sold by retailers. However, they also found that these outbreaks, especially those associated with lettuce, tended to be larger than those caused by other food types and more prolonged. There has also been some analysis of foodborne disease outbreaks in the USA between 1973 and 2005 and this shows that "fresh plant produce" was responsible for only 0.7% of outbreaks in the 1970s, but that this had risen to 6% by the 1990s. While some of this increase can be accounted for by rising consumption, the figures show that, between 1986 and 1995, the number of foodborne disease outbreaks linked specifically to leafy greens rose by 60%, but consumption rose by only 17%. The trend from 1995 to 2005 was similar, pointing to a growing problem with the safety of vegetables like lettuce and spinach.
Prepared salads in general are clearly not a major contributor to foodborne illness, but on both sides of the Atlantic there is evidence of a disproportionate problem with foodborne disease linked specifically to leafy green salad vegetables, especially lettuce and spinach.
A global perspective
Concern over the safety of leafy greens is growing at an international level. For example, a joint WHO FAO expert meeting on microbiological hazards associated with fresh produce in October 2007 concluded that leafy green vegetables should be given the highest priority in terms of fresh produce safety. There were a number of reasons for this, but the assembled experts considered that, "from a global perspective," this group of foods gave rise to the greatest concern because of their potential to cause large and widespread foodborne disease outbreaks and the potential for post-harvest processing to "amplify" contamination.
Why were the experts at that meeting so concerned? One reason is a worrying trend towards more frequent multinational foodborne disease outbreaks linked to green salad vegetables, especially lettuce. A typical example was reported in 2000, when at least 140 people in Denmark, Germany, Iceland, the Netherlands and the UK were infected by a strain of
These examples illustrate part of the problem. The demands of consumers for fresh salad greens all year round mean that lettuce and other vegetables have to be sourced from many producer countries, creating highly complex supply chains. Produce from a single grower can end up in several countries and can be supplied to many customers. If a pathogen enters this system it can be spread over a wide geographical area very quickly and can be very difficult to trace. Food safety experts are beginning to realise that the global trade in leafy salad greens, driven by consumer demand for healthy eating, may in fact be an important vehicle for spreading human pathogens around the world.
Salmonella typhimurium. The outbreak was associated with lettuce, but the supply chain was so complex that its point of origin was never confirmed. In another incident in 2007 at least 50 people in Iceland and the Netherlands became ill with E. coli O157 infection after consuming shredded, pre-packed lettuce from a Dutch processing plant. Other leafy greens and fresh herbs have also been involved in outbreaks, such as rocket lettuce from Italy and fresh basil grown in Israel.
How contamination occurs
It is easy to come up with a list of possible sources of pathogen contamination for crops growing in the field: organic manure added to the soil; domestic and wild animals; birds; water contaminated with animal or human faecal material; poor hygiene by agricultural workers. What is much more difficult is tracing contaminated salad greens back to an individual grower and establishing the exact source of contamination.
For instance, an outbreak of Salmonella senftenberg in 2007, which caused at least 50 cases in the UK, the Netherlands, Denmark and the USA, was linked to fresh pre-packed basil grown in Israel. Exhaustive environmental investigations in Israel failed to reveal the source of the contamination. A similar situation occurred with a 2004 outbreak of Salmonella thompson infection, which caused 100 cases in Denmark, Norway, Sweden and the UK and was linked to rocket lettuce grown in Italy. On this occasion investigators suspected sewage-contaminated irrigation water as the source, but this could not be confirmed. Investigators had more luck when looking into an outbreak of E. coli O157 in Norway and Sweden in 2005. This outbreak was associated with consumption of iceberg lettuce, this time grown locally in Sweden, allowing a more rapid and thorough investigation. The source was identified as a small stream used to irrigate the crop. Perhaps the most exhaustively investigated event was the high profile spinach-related E. coli O157:H7 outbreak in the USA in 2006. The spinach was traced back to farms in the Salinas Valley in California, the source of at least 10 other outbreaks since 1995. The outbreak strain of E. coli O157:H7 was isolated from a stream, cattle manure and the faeces of wild pigs in the area. This suggested that the growing environment could become contaminated from more than one source, with the pathogen being dispersed by water and by the movement of animals. The findings present a more complex picture than anyone expected and show the need for more research into the sources and movement of pathogens in the field.
Control and decontamination options
So how can contamination of salad greens with pathogens be prevented and/or controlled? Unfortunately there is no simple answer. The most productive approach at the growing stage is based on the established principles of Good Agricultural Practice (GAP). GAP is intended to help farmers minimise environmental pollution and protect natural resources. These general principles have been used as the basis for developing codes of practice for growers. For example, in the USA the FDA Center for Food Safety and Applied Nutrition has published a Guide to Minimize Microbial Food Safety Hazards for Fresh Fruits and Vegetables. It contains advice on key elements for preventing contamination, including clean water supplies, safe use of manure, agricultural worker health and hygiene, hygienic practice in the field, in packing and in transport, and traceability. But such guidance can only achieve so much. Any crop growing in the field is vulnerable to contamination from sources not under the growers’ control, such as wild animals and birds. To ensure absolute safety, post-harvest treatments are needed.
Unfortunately, completely effective post-harvest methods for treating fresh produce to eliminate microbial pathogens without destroying the sensory qualities of the product are simply not available. Traditionally, the main decontamination method available to the produce industry has been washing with potable water, often chlorinated. Washing with chlorinated water is routinely applied to leafy green vegetables like lettuce, especially for pre-packed ready-to-eat salads, but its effectiveness is limited. For example, the investigation into the 2006 US
This has led some food safety experts to back rather more drastic decontamination methods, notably irradiation. Researchers have shown recently that a relatively low dose of 1 kGy – enough to give a theoretical 5-log reduction in
E. coli O157 outbreak linked to spinach found that the spinach had been washed prior to packing and that further washing by the consumer made no difference to the chances of becoming infected. Washing rarely achieves microbial reductions of more than 100-fold even when 100 ppm or more of free chlorine is present. Other sanitisers have been tried, including chlorine dioxide and ozonated water, but all have their disadvantages. E. coli O157 – did not damage the quality, or nutritional value, of fresh cut vegetables, including lettuce. In fact, the treatment actually delayed spoilage to some extent. In the USA, the FDA announced last year that it was proposing to allow irradiation of fresh spinach and iceberg lettuce with an absorbed dose of up to 4 kGy to control pathogens. Given consumer attitudes to food irradiation, it is doubtful whether the process represents a practical solution to the problem of pathogens in green salad vegetables. Some microbiologists believe that the best solutions lie in interventions that can be applied in the field, rather than in better decontamination processes, and they point to some very recent research, which may reveal new ways to attack pathogens before they become established in fresh produce.
New insights may lead to new controls
While we are beginning to identify the main sources of contamination for crops growing in the field, the mechanisms by which human pathogens attach themselves to plants are much less obvious. If contamination were a purely passive process one might expect that decontamination by washing lettuce and other crops with chlorinated water would be effective. The evidence suggests that this is not the case and that other factors are involved. The interaction between human pathogens and plants is a relatively new field of research, but some interesting findings have already been published and the emerging picture seems to be surprisingly complex.
So far the evidence indicates that contamination of food plants by human pathogens is more likely to occur in the phyllosphere (the above ground plant surfaces) rather than via the root systems. This seems to be especially so for lettuce. There is also evidence that both
Some researchers have now begun to look more closely at bacterial attachment in the phyllospere of salad greens. For example, a team from Imperial College in the UK looked at
This research is still in its infancy, but if we can learn more about these attachment mechanisms and how they work, we may be able to gain a better understanding of the risk factors involved and perhaps devise controls to inhibit attachment in the field. Perhaps it will be possible to breed crop varieties that are resistant to pathogen attachment, or to spray crops with compounds that inhibit the attachment process. What seems certain is that a lot more research will be needed before we can solve the problem of contaminated salad crops. We may just have to be content with minimising the risk through GAP and good hygiene until then. That being the case, it is fortunate that salad-related food poisoning outbreaks are still comparatively rare.
Salmonella and E. coli can become internalised within leafy tissue, either through damaged areas, via cut surfaces produced during harvesting, or by entering the plant through the stomata. For instance, E. coli O157:H7 cells have been shown to be attracted to the guard cells surrounding the stomata on leaf surfaces. Human pathogens may also become components in microbial biofilms attached to leaf surfaces and this may help to attach the cells more firmly and protect them from washing and sanitising chemicals. There is even evidence that some pathogens can multiply on leaf surfaces. A US study published in 2008 showed that, given warmth and moisture, both E. coli O157:H7 and Salmonella enterica could multiply by up to 100-fold in the phyllosphere of young lettuce plants and on the harvested leaves of older plants. Growth of human pathogens inside leaf tissues has not yet been conclusively demonstrated, but this too is a possibility.E. coli types O157 and O26 and were able to identify how they were attached. These bacteria seem to use structures called EspA filaments projecting from the cell wall to attach to leaf surfaces. The same structures are an important part of the mechanism that these pathogens use to infect the cells lining the human gut and the research suggests the intriguing possibility that these bacteria use the same method to attach to both mammalian and plant cells, although they don’t cause disease in the plant. The same team have also looked at the strain of Salmonella senftenberg associated with the outbreak linked to Israeli-grown fresh basil and investigated how it was able to attach to the leaf surfaces. They found that the bacteria were able to bind to basil, lettuce, rocket and spinach leaves and they reported that flagella could be seen linking the bacterial cells to the leaf surface. Deleting the gene responsible for flagella production reduced the amount of attachment. But Salmonella typhimurium did not behave in the same way – it was not affected by deletion of the same gene – and the researchers suggest that different Salmonella strains could use different leaf attachment mechanisms. They also reported that the leaves of some plants were much less susceptible to Salmonella contamination than others.
References
Grant J.
Little C L. & Gillespie I A. Prepared salads and public health. Journal of Applied Microbiology 2008; 105: 1729-43.
et al. Spinach-associated Escherichia coli O157:H7 outbreak, Utah and New Mexico, 2006. Emerging Infectious Diseases 2008; 14(10).
