Lessons Learned from a Decade of Investigations of Shiga Toxin-Producing Escherichia coli Outbreaks Linked to Leafy Greens, United States and Canada.

Shiga toxin-producing Escherichia coli (STEC) cause substantial and costly illnesses. Leafy greens are the second most common source of foodborne STEC O157 outbreaks. We examined STEC outbreaks linked to leafy greens during 2009-2018 in the United States and Canada. We identified 40 outbreaks, 1,212 illnesses, 77 cases of hemolytic uremic syndrome, and 8 deaths. More outbreaks were linked to romaine lettuce (54%) than to any other type of leafy green. More outbreaks occurred in the fall (45%) and spring (28%) than in other seasons. Barriers in epidemiologic and traceback investigations complicated identification of the ultimate outbreak source. Research on the seasonality of leafy green outbreaks and vulnerability to STEC contamination and bacterial survival dynamics by leafy green type are warranted. Improvements in traceability of leafy greens are also needed. Federal and state health partners, researchers, the leafy green industry, and retailers can work together on interventions to reduce STEC contamination.

A systems analysis of irrigation water quality in an environmental assessment of an E. coli O157:H7 outbreak in the United States linked to iceberg lettuce.

A foodborne Escherichia coli O157:H7 outbreak in December 2006 included 77 illnesses reported in Iowa and Minnesota. Epidemiologic investigations by health departments in those states and the U.S. Centers for Disease Control and Prevention (CDC) identified shredded iceberg lettuce (Lactuca sativa L.) as the vehicle of transmission. The U.S. Food and Drug Administration (FDA) and Minnesota and California public health agencies traced the lettuce to several growing regions in California based on information from a lettuce processor in Minnesota. Samples from an environmental investigation initiated by the California Food Emergency Response Team (CalFERT) revealed a genetic match between the outbreak strain and environmental samples from a single farm, leading to an in-depth systems-based analysis of the irrigation water system on that farm. This paper presents findings from that systems-based analysis, which assessed conditions on the farm potentially contributing to contamination of the lettuce. The farm had three sources of irrigation water: groundwater from onsite wells, surface water delivered by a water management agency and effluent from wastewater lagoons on nearby dairy farms. Wastewater effluent was blended with the other sources and used only to irrigate animal feed crops. However, water management on the farm, including control of wastewater blending, appeared to create potential for cross-contamination. Pressure gradients and lack of backflow measures in the irrigation system might have created conditions for cross-contamination of water used to irrigate lettuce. The irrigation network on the farm had evolved over time to meet various needs, without an overall analysis of how that evolution potentially created vulnerabilities to contamination of irrigation water. The type of systems analysis described here is one method for helping to ensure that such vulnerabilities are identified and addressed. A preventive, risk-based management approach, such as the Water Safety Plan process for drinking water, may also be useful in managing irrigation water quality.

Overview of Leafy Greens-Related Food Safety Incidents with a California Link: 1996 to 2016.

An increase in the number of foodborne illness outbreaks associated with produce has been noted in the literature, and leafy greens have been the most common produce category associated with these outbreaks. California is the largest leafy greens producer in the United States, and many related foodborne illness incidents were traced to this state. A systematic overview of leafy greens incidents linked to California was conducted by the California Department of Public Health, Food and Drug Branch through analysis of complaints, routine surveillance sampling, disease outbreaks, and investigations covering 1996 to 2016. The goal was to develop a risk assessment tool to modernize emergency response efforts to foodborne illnesses related to leafy greens. A database including environmental, epidemiologic, and laboratory information for each incident was developed, and descriptive analysis was performed to identify trends. In the 21-year period analyzed, 134 incidents were identified, the majority of which were surveillance related. Approximately 2,240 U.S. cases of confirmed illness were reported (298 California cases resulting in 50 hospitalizations). Romaine lettuce and spinach were the most commonly implicated vehicles. The most prevalent hazard type was microbiological, in particular bacterial, specifically associated with pathogenic strains of Escherichia coli. In California, the overview provided the Food and Drug Branch with a platform to (i) enhance its Food Safety Program, Emergency Response Unit, and California Food Emergency Response Team; (ii) assist in more efficient investigation, response, control, and prevention of California-linked foodborne illness incidents; and (iii) identify knowledge gaps and develop effective definitions, procedures, training, guidelines, and policies that will be used to help prevent future outbreaks. Outcomes provide insight into the situation in the largest leafy greens-producing state and may be used to prioritize limited national food safety resources and aid in future leafy greens-related research and foodborne incident investigations.

Ammonia disinfection of animal feeds --laboratory study.

Animal feeds may be contaminated, accidentally or maliciously, with a number of zoonotic bacteria. Animal infections with these bacterial agents, whether or not they cause animal disease, may lead to human illnesses. Anhydrous ammonia was introduced on farms in developed countries as a high-nitrogen soil amendment, but later found use in enhancing crude protein in low-quality roughage fed to ruminants and in neutralizing mycotoxins in fungus-infested feed grains. Although ammonia has been known to be effective against bacteria in other contexts (e.g., manure, community sewage sludge, seeds for sprouting, and boneless lean beef trimmings), it appears that the antibacterial effect of ammoniating animal feeds had not been tested. In the present study, samples of roughage (wheat straw, corn silage) and concentrates (corn grain, cottonseed) produced as animal feed were contaminated with dried-on zoonotic bacteria (Salmonella Newport in all; Campylobacte jejuni, E. coli O157:H7, Listeria monocytogenes, Yersinia enterocolitica in corn grain only). Disinfection with anhydrous ammonia gas was conducted for 24 h at room temperature ( 25 degrees C). The treatment was least effective in silage because the silage alone showed strong antibacterial activity, which may have been slightly reduced by ammoniation. In the other three feeds, depending on the initial level of contamination, ammonia destruction of >or= 5 log10 cfu/g (99.999%) of the selected contaminant was usually observed.

Pathogen survival in chorizos: ecological factors.

This study addressed health risks from ethnic sausages produced on a small scale, without inspection, in California and elsewhere. Mexican-style chorizo, a raw pork sausage that is not cured, fermented, or smoked, was contaminated experimentally in the batter with Escherichia coli O157:H7, Listeria monocytogenes, or Salmonella serotypes and stuffed into natural casings. Formulations were based on a market survey in California. Physical parameters that were controlled were pH, water activity (a(w)), and storage temperature. The pH was adjusted with vinegar, stabilizing at 5.0 within 24 h. Initial a(w) levels adjusted with salt were 0.97, 0.95, 0.93, 0.90, and 0.85; levels declined with time because of evaporation. Pathogen numbers declined with storage up to 7 days, with few brief exceptions. Main effects and interactions of constant temperature and pH with declining a(w) on survival of the pathogens were determined. Maximum death rates occurred at higher a(w) for E. coli O157:H7 and Salmonella than for L. monocytogenes. Salt used to adjust a(w) affected palatability. Spices (black pepper, chili pepper, chili powder, cumin, garlic, guajillo pepper, oregano, and paprika) comprised another, potentially significant aspect of the sausage formulation. Some (notably black pepper and cumin) carried an indigenous microflora that contributed significantly to the microbial load of the sausage batter. Only undiluted fresh and powdered garlic exhibited a significant antimicrobial effect on the pathogens. Although each of the tested formulations caused death of the inoculated pathogens, none of the death rates was sufficiently rapid to ensure safety within the probable shelf life of the product.

Central nervous system tissue detection in meat from advanced meat recovery systems.

Three hundred meat samples, recovered from beef neck- and breast-bones using a conventional advanced meat recovery (AMR) system, the de-sinewed minced meat (DMM10) technology, and hand-boning, were collected and tested for presence of central nervous system tissue (CNST) in meat using an ELISA-based test. Samples were collected at two processing facilities (Est. A and B). Sternum meat was the non-CNST reference (control) - it is distant from brain and spinal cord locations on a carcass, with low likelihood of contamination with CNST. Neckbone meat was recovered from bones obtained from carcasses where the spinal cord was removed manually, Est. B, or using a Jarvis circular hydraulic cord remover saw, Est. A. All samples from AMR, DMM, and hand methods showed lower calculated levels of "risk material" than the stated limit of detection (0.1%) of ELISA kit. There was no apparent difference among these, and use of the Jarvis saw had no perceptible advantage.

Survival of Listeria monocytogenes in experimental chorizos.

Chorizos-Mexican-style raw-meat sausages-are a concern in California because their production in small ethnic food markets is unregulated. Their formulation may cause them to appear cooked to the consumer, who may eat the raw sausage without prior proper cooking. Bacterial pathogens in such products may cause illness or even death. Survivability of Listeria monocytogenes in chorizos was evaluated under different storage conditions selected on the basis of an initial survey of uninspected chorizos in California. Sausages were formulated to five different initial water activity (aw) levels (0.85, 0.90, 0.93, 0.95, 0.97), stored under four conditions (refrigeration, "Ref," 6 to 8 degrees C under convective air circulation; room temperature, "RT," 24 to 26 degrees C under convective air circulation; hood, "Hd," 24 to 26 degrees C under forced air circulation; and incubation, "Inc," 30 to 31 degrees C under convective air circulation), and sampled after 1, 2, 4, and 7 days. The initial pH was 4.8 and remained near 5.0 from day 1 of the sampling period. An inoculated-pack study using a five-strain cocktail of L. monocytogenes was performed twice for each initial aw. Results indicated that the three lowest initial aw levels (0.85, 0.90, 0.93) and the Hd and Inc storage conditions were more effective (P < or = 0.05) at reducing L. monocytogenes levels in chorizos than the two highest initial aw levels (0.95 and 0.97) and the Ref storage condition, irrespective of storage time. These results can provide a scientific basis for guidelines given to uninspected chorizo producers in California and reduce the risk of foodborne illness.

Survival of Salmonella and Escherichia coli O157:H7 in chorizos.

Mexican-style raw meat sausages (chorizos) are not regulated in California when they are produced in small ethnic food markets. These sausages are sold uncooked, but their formulation imparts a color that may lead the consumer to assume that they are already cooked, and thus the chorizos may sometimes be eaten without proper cooking. If pathogens are present in such cases, illness may result. Survival of Salmonella and Escherichia coli O157:H7 in chorizos was evaluated under different storage conditions selected based on an initial survey of uninspected chorizos in California. Chorizos were formulated with five different initial water activity (aw) values (0.85, 0.90, 0.93, 0.95, and 0.97), stored under four conditions (refrigeration at 6 to 8 degrees C, room temperature at 24 to 26 degrees C, under a hood at 24 to 26 degrees C with forced air circulation, and incubation at 30 to 31 degrees C with convective air circulation), and sampled after 1, 2, 4, and 7 days. The initial pH was 4.8 and remained near 5.0 from day 1 of the sampling period. Two separate studies of packs inoculated with five-strain cocktails of Salmonella and of E. coli O157:H7 were performed twice for each initial aw. The three lowest aw values (0.85, 0.90, and 0.93) and the incubation and hood storage conditions were more effective (P < or = 0.05) at reducing the target pathogen levels in chorizos than were the two highest aw values (0.95 and 0.97) and the refrigeration storage condition, regardless of storage time. These results provide a scientific basis for guidelines given to producers of uninspected chorizo and should reduce the probability of foodborne illness associated with these products.

Spinal cord tissue detection in comminuted beef: comparison of two immunological methods.

Two commercial immunological kits for detection of central nervous system (CNS) tissue in beef were compared: ScheBo® Brainostic™, based on CNS-specific antigen (neuron specific enolase) detection, and Ridascreen® Risk Material 10/5 test, an enzyme immunoassay for glial fibrillary acidic protein. Spinal cord (SC) was added to batches of choice, select, and utility grades of ground fresh beef shoulder clod to yield 0.0, 0.0125, 0.025, 0.05, 0.1, 0.2, 0.4, 0.8, and 1.6% SC in meat. Sensitivity and specificity in detecting SC in fresh and frozen samples were determined. Both Brainostic™ and Ridascreen® kits detected SC at claimed levels: 0.25% and 0.11%, respectively. The Ridascreen® test consistently detected SC at 0.025%, below its claimed sensitivity level, expressed for brain and SC combined. The Ridascreen® test was ∼10× more sensitive, easier, faster to run and less expensive than the Brainostic™. Overall, quality grade had no influence on SC detection in fresh or frozen meat.

Impact of sodium chloride on Escherichia coli O157:H7 and Staphylococcus aureus analysed using transmission electron microscopy.

Abundant literature information is available on sodium chloride, NaCl, as an antimicrobial and a preservative, however, information on NaCl effects on bacterial cell morphology is lacking. The effect of NaCl, on Escherichia coli O157:H7 and Staphylococcus aureus cells individually grown in a laboratory medium was examined using transmission electron microscopy (TEM). Cultures were grown in brain heart infusion (BHI) broth containing dissolved 0%, 5%, or 10% (w/v) commercially obtained fine (FN) and extra coarse (EC) grade granular NaCl. The pathogens were incubated at 35 degrees C for 12 and 24 h. Then, a mixture of five strains of each pathogen per treatment was prepared. Samples were centrifuged, pellets collected, fixed immediately with glutaraldehyde, and prepared for TEM examination. Cells morphology on TEM micrographs verified that the magnitude of morphological damage to E. coli O157:H7 cells was significantly greater than that of S. aureus cells. More cell injury occurred as NaCl concentration increased from 5% to 10%. Generally, S. aureus maintained its cellular structure and no severe cell wall or plasma membrane damage and/or shrinkage was observed. At 10% NaCl, the damage to E. coli O157:H7 cells was extensive, and the pathogen seemed to have lost its cellular integrity. Although NaCl affected the morphology of E. coli O157:H7 and S. aureus, the coarse grade of NaCl seemed to have a milder effect with respect to cell damage, especially on S. aureus. The 24 h-old cultures were more susceptible to NaCl treatment compared to the 12 h-old cells. Thus, the age of the cells has an impact on their resistance to salt--the environmental stressor.