Although we’re advised to wash our fresh fruits and veggies before eating them, this procedure only deals with surface contamination. Produce, such as salad vegetables and fruits that are usually consumed raw, can still be contaminated with human pathogens such as Salmonella species and E.coli internalized within their cells. Contamination can come from the soil, water supply, manure/fertilizer or human contact and once inside the cell, the bacteria cannot simply be washed away even if a chemical treatment is used. Efficient and sensitive detection is vital for maintaining food safety. Although bacterial isolation definitively shows contamination, it is a slower and less sensitive method than newer molecular-based strategies such as polymerase chain reaction (PCR) assays. Although contamination can occur at any stage in the food production cycle, pathogen internalization usually happens while the plant is growing. Environmental conditions such as water stress or disease can affect this process. Ge et al.1recently investigated the effect of abiotic stress (too much or too little water) and biotic stress (infection with lettuce mosaic virus) in growing lettuce. Using green fluorescence protein-labelled Salmonella enterica serovar Typhimurium, they contaminated the leaf surfaces of the growing plants then examined the effects of the two stressors on bacterial internalization. After surface decontamination, confocal microscopy showed that the Salmonella species were indeed internalized in the leaves and root systems of contaminated lettuce plants. Moreover, colony plate counts showed that viable bacteria were present in the leaves and root systems of water-stressed plants infected with the mosaic virus. Interestingly, only PCR showed the presence of pathogen DNA in the roots of uninfected, water-stressed plants. Ready-to-eat salads and fresh fruits are popular diet choices as people adopt healthier lifestyle. Since timely harvesting and delivery to the consumer is vital for absolute freshness, food safety testing needs to be rapid, efficient and sensitive. Results are needed swiftly for a timely response to protect human health. One solution to this problem is the Thermo Scientific SureTect™ Salmonella species quantitative real time PCR (qRT-PCR) assay. This probe-based kit has recently been granted Performance Tested Method(SM) status by the AOAC Research Institute and is comparable to the reference method ISO 6579:2002, “Microbiology of food and animal feeding stuffs: Horizontal method for the detection of Salmonella species”. The assay offers a convenient way to analyze multiple samples specifically for the invA gene found only in this pathogen. One of the main problems in PCR analysis of food matrices is the presence of interfering factors and the requirement for sample enrichment. These are dealt with in a simple one-step sample preparation step using pre-filled lysis tubes at the beginning of the SureTect assay. All reagents are provided and the dedicated software gives easily interpreted results on completion. Reference
1. Ge, C. et al. (2014) “Impact of phytopathogen infection and extreme weather stress on internalization of Salmonella Typhimurium in lettuce,” International Journal of Food Microbiology 168–169(pp.24–31)
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We now have the capability to find and measure minute levels of contamination. While I understand it is important to know if a risk exists, The risk then needs to be characrized. How do we propose to do that?
Hi Jim – this is a great question. This article mentions a rapid method for the detection of salmonella in produce. In the U.S., if we were to get a positive result for salmonella on a rapid method, we would have to follow the guidelines set by the FDA in the Bacterial Analytical Manual, or FDA BAM. For this specific example, the potentially contaminated lettuce would be incubated in lactose broth for 24 hours. From here, depending on the microbial load of the lactose broth incubation, subsequent incubations in various other media types under different incubation conditions would take place. For more detailed information on the characterization of salmonella from different food matricies, please visit http://www.fda.gov/Food/FoodScienceResearch/LaboratoryMethods/ucm070149.htm#Isol, or feel free to leave us another comment!
I see how to do the cultures for confirmation. What I do not see is the level at which the FDA believes or prescribes intervention. How is the risk calculation done? Also, what is the intervention and how is the excessively contaminated product dealt with? Can it be cooked? Can it be irradiated?
My understanding is the FDA is considering defining some Salmonella species as adulterants. In the produce world, the organism may lodge in tubules. Does it colonize and proliferate? In pigs, Salmonella colonizes lymph nodes. All lymph nodes cannot be removed, so there will always be a risk that individual lots of pork will be “adulterated” and ineligible for sale. I am unsure if cooking makes it eligible for sale.
Hi Jim – the potential risk is determined based on the
contaminant or adulterant, and the food matrix it is associated with. For example, Ethylene Dibromide has an action
level of 0.3 ppb in fish for the edible portion, and 0.01 ppb for leafy vegetables, excluding brassica. For more info, please see the following link http://goo.gl/vkwWRp. It is also important to note the difference between a contaminant and an adulterant for readers who may be unaware of the distinction. A contaminant is broadly deemed any substance that is foreign to the food product, while an adulterant is defined in six different ways that could lead to criminal charges of negligence, including fines and imprisonment, for knowingly introducing these foods into commerce. Adulterants are defined in the Code of Federal Regulations under 9 CFR 301.2, which can be found here http://goo.gl/Brj0cj. Foods that are sold raw but are intended to consume cooked, such as poultry and eggs, are potential carriers of pathogens that may cause foodborne illness. As long as these foods are cooked properly, this greatly reduces the chance of foodborne illness caused by contaminants. Under the current regulations set forth by the FDA and the Food Safety and Inspection Service (FSIS), any food product containing an adulterant is unsafe for sale and consumption, regardless of any measures taken on behalf of the consumer. The bill I believe you are referring to is S. 1529, which aims to include antibiotic-resistant Salmonella serovariants, as well as Shiga toxin-producing E. coli. I will be following this legislation as well due to the impact it will have on the food industry. Another great question, Jim!
I think characterizing the risk would take a lot of experimentation, and each regulatory body must make a judgement based on the best evidence available. Again, GMP/GHP from farm to fork should be observed as basic standard practice in HACCP. All this helps to minimise the risk. Sometimes, additional practices/control mechanisms may be employed with partial risk characterization. Any comments?
How do we interpret results from DNA tests in identifying and quantifying live cells?
Hi Beatrice – another great question! Through real-time PCR, it is possible to quantify gene copy numbers, which can then be traced back to the number of cells or colony forming units (cfu’s) when proper controls are used. Real-time PCR is a very sensitive and specific method to detect DNA from living, growth-inhibited, and dead cells by targeting unique segments in the gene of interest. Because real-time PCR detects DNA and not just living cells, it is emerging as a popular screening method for the rapid identification for the presence of pathogens. If a positive result is obtained, then the pathogen must be further characterized by serology or culture methods as perscribed by whichever regulatory body has jurisdiction over the food lab. For an example of the results obtained by the SureTect real-time PCR system, please see the attached picture. If you have any other questions, please let us know!