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  1. Sampling to Protect the Food Supply

Microbiological Surveillance Sampling: FY20-21 Sample Collection and Analysis of Romaine Lettuce (Raw Agricultural Commodity)

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The FDA conducted a small, focused assignment in 2019 and 2020 to collect samples of raw agricultural commodity romaine lettuce to test for pathogenic Escherichia coli (specifically, Shiga toxin-producing E. coli or STEC) and Salmonella spp., microbial hazards repeatedly linked to foodborne illnesses associated with romaine lettuce consumption. The FDA did not detect any pathogens during the assignment, which ended in December 2020.

The FDA began the assignment in November 2019, following multiple outbreaks of foodborne illness linked to romaine lettuce. The United States experienced two multistate outbreaks of E. coli O157:H7 infections in 2018 (in the spring and fall), and four outbreaks of E. coli O157:H7 infections in 2019 (one in the summer, and three in the fall), all linked to or possibly linked to the consumption of romaine lettuce. In the fall of 2020, the FDA investigated a multistate illness outbreak linked to leafy greens and caused by a strain of E. coli genetically related to a strain associated with the 2019 fall romaine lettuce outbreaks. Salmonella spp. also commonly cause foodborne illness outbreaks associated with fresh produce consumption in the United States, and in 2013 the nation experienced a multistate outbreak of Salmonella Newport infections linked to romaine lettuce consumption.

The FDA’s goal in conducting this assignment was to see whether the target pathogens may be present at the FDA-registered facilities and farms linked by FDA traceback investigations to foodborne illness outbreaks of recent years where romaine lettuce was confirmed or suspected to be the food vehicle. The agency collected most of the samples at facilities in the Yuma, Arizona, and Salinas Valley, California, agricultural regions. In the event that STEC or Salmonella spp. were detected, the FDA intended to conduct follow-up inspections to evaluate controls and determine potential sources and routes that may have led to the contamination. Although the FDA did not detect any pathogens during this assignment, maintaining surveillance during the leafy greens growing/harvesting season is helpful to informing the agency’s leafy greens prevention efforts.


The FDA set out to collect and test 270 samples of romaine lettuce in November 2019 for this assignment. The agency collected and tested a total of 279 samples for both pathogens. Each sample was made up of 10 subsamples, with each subsample consisting of one or more heads or hearts of romaine lettuce and weighing at least 300 grams. This approach – the  collection and testing of samples composed of multiple subsamples – increases the probability of detecting pathogens if present, given that microbial hazards may not be uniformly present. Agency field staff collected all samples in their natural form but for the outer leaves having been removed. No fresh-cut lettuce was collected.

The agency prioritized sample collection at FDA-registered facilities and farms identified in traceback investigations as possible suppliers of romaine lettuce linked to outbreaks of foodborne illnesses from 2017 to 2019. FDA investigators collected samples from wholesalers, distribution centers, on-farm holding locations, and commercial cooling and cold storage facilities, and to a lesser extent, from refrigerated storage, prior to consumer access at grocery stores. No samples were collected from farm fields.

The FDA designed its assignment to emphasize collection of samples early in the distribution chain. Sampling of raw agricultural commodity romaine lettuce before it is commingled during fresh-cut processing or before preparation at point of service, where it may be commingled with other produce, would facilitate traceback of the lettuce had samples tested positive for human pathogens.

In testing for STEC, FDA analysts followed the methods in Chapter 4A of the agency’s Bacteriological Analytical Manual (BAM). In testing for Salmonella spp., the analysts followed the methods in BAM Chapter 5. The FDA also planned to conduct whole genome sequencing (WGS) of any pathogens detected to identify their genetic patterns and determine whether those pathogens may be linked to human illness.

COVID-19 and Sample Integrity

Five months into this assignment (in mid-March 2020), the agency paused food product surveillance sampling in response to the COVID-19 pandemic. This assignment remained on hiatus through the summer, along with others, as the FDA evaluated operational activities and prioritized those it deemed to be mission critical. The agency designated this field activity as mission critical this past fall, coinciding with the most recent multistate outbreak of E. coli O157:H7 infections associated with leafy greens. In October 2020, the agency resumed sample collection and testing to complete this assignment.

Agency field staff collected all samples aseptically to prevent contamination during the collection process, in accordance with the FDA’s Investigations Operation Manual (IOM). The FDA’s aseptic sampling methods, which entail the use of sterile implements and containers, and prescribed collection procedures, are published in IOM chapter 4.3.

The FDA took additional precautions given the COVID-19 pandemic when it resumed sample collection. Agency field staff were outfitted with personal protective equipment and observed all local, state and applicable Centers for Disease Control and Prevention guidance.


All the samples collected and tested for this assignment were negative for Salmonella spp. and strains of E. coli capable of causing serious illness. The agency detected E. coli in one sample, but subsequent characterization showed the organism did not feature any known characteristics that would enable it to adhere to intestinal epithelium, which is essential for infection to begin. The FDA also conducted WGS on the organism and determined that it was not associated with any known human illnesses.


The findings of this assignment suggest that microbial contamination – to the extent that it may have been present – was not widespread at the 137 FDA-registered facilities and farms where agency investigators collected samples during the period of this field activity. The FDA cautions against making any further inferences based solely on this assignment’s findings given that the sample size was relatively small and in view of the fact that multiple foodborne illness outbreaks linked to romaine lettuce have occurred in recent years.

The agency is working on several fronts to help prevent microbial contamination in romaine lettuce and other leafy greens, which are among the most widely consumed vegetables and an important part of a healthy diet. Chief among the FDA’s efforts is its 2020 Leafy Greens STEC Action Plan.

The FDA published and implemented its action plan last year and will continue to work with stakeholders throughout the supply chain to strengthen the microbiological safety of leafy greens. While some of the FDA’s efforts have been constrained by the COVID-19 pandemic, the agency continues to make progress toward the plan’s objectives. The plan features public health and regulatory approaches in three categories: response, prevention and the addressing of knowledge gaps.

Research is a critical part of the plan, as the FDA has identified key questions to address and answer to inform prevention efforts, with the ultimate goal of improving the microbiological safety of leafy greens. To that end, the FDA has launched two longitudinal studies based in the lettuce growing regions of Arizona and California, respectively, to better understand how human pathogens survive and migrate through the environment in those regions. These multi-year studies center on the collection and testing of environmental samples, including adjacent land, well and surface waters, and soil inputs that may contain compost, dust and animal fecal material. The studies are being conducted in partnership with state and local governments and universities; they are intended to strengthen prevention efforts by informing agricultural practices in those regions.

Consumers can reduce risk and thereby help protect themselves and their families by thoroughly washing romaine lettuce and other leafy greens under running water before eating them. Washing leafy greens is not a lethality step (i.e., it is not guaranteed to eliminate pathogens) but may well remove some pathogens, if present. Many pre-cut, bagged, or packaged produce items are pre-washed and ready-to-eat. If so, it will be stated on the packaging, and consumers can use the produce without further washing. Consumers who choose to wash leafy greens marked as “pre-washed” or “ready-to-eat” should make sure that it does not come in contact with unclean surfaces or utensils. This will help to avoid cross contamination. Additional information on leafy greens and food safety is available at the website of the U.S. Centers for Disease Control and Prevention. Romaine lettuce and other leafy greens require appropriate protection from human pathogens during growing, harvesting, packing, processing and holding – in short, during all stages from farm to table.

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