There are multiple detection methods for Salmonella. Traditional cultural methods for isolation include plating on selective agars and incubating for 24 h at 35 °C . Before plating, naturally-contaminated samples are often enriched with non-selective and/or selective broths such as lactose broth since a low concentration of Salmonella is expected. Standard methods and media used to isolate Salmonella can be found in the FDA Bacteriological Analysis Manual . In conjunction with traditional cultural methods, rapid biochemical or antigen-antibody-based methods can be used for quicker isolation and identification of Salmonella . Salmonella can also be identified through testing a combination of biochemical and serological reactions. Most Salmonella will provide a positive result for glucose , lysine decarboxylase , H2S , lysine carboxylase broth, phenol red dulcitol broth, polyvalent flagellar test, polyvalent somatic test, and methyl red test; and provide a negative result for urease, potassium cyanide broth, malonate broth, indole test, phenol red lactose broth, phenol red sucrose broth, and VogesProskauer test Salmonella can be further identified through phenotyping methods such as serotyping, phage typing, biotyping, and R typing . Finally, Salmonella can be identified through genotyping by PCR or pulse-field gel electrophoresis . PFGE was a highly used method to trace outbreaks, but whole genome sequencing is now the current method used by PulseNet . Whole genome sequencing is a laboratory procedure that determines the order of bases in the genome of an organism in one process . Because millions of bases make up the WGS for every organism, grow bag for tomato it is much more detailed method than Pulse Field Gel Electrophoresis which was the former gold standard method for differentiating among pathogen isolates .

The CDC started implementing the use of WGS as its main way tracking foodborne outbreaks in 2013 . They are able to compare genomes from outbreak strains to reference genomes from public data bases such as EnteroBase . Shiga toxin producing Escherichia coli. Shiga toxin producing E. coli is a gramnegative, non-spore-forming bacteria that can cause infection in humans. Like Salmonella, it belongs to the family Enterobacteriaceae. STEC can grow in temperatures ranging from 7 °C to 45 °C but has optimal growth from 35 °C to 42 °C . It can grow in a pH range of 4-10, and requires a water activity of 0.95 or higher . STEC can be carried by many types of animals and is commonly associated with ruminants such as cattle . STEC will be passive in many of these hosts, but can cause disease in humans. Symptoms of infection by STEC include bloody diarrhea, vomiting, and in certain cases hemolytic uremic syndrome . STEC infects humans by using attachment and effacement lesions encoded for on their LEE pathogenicity island . As the name suggests, the main toxins used by STEC are Shiga toxins, which is what leads to cell death in the host. Apart from being the most known disease-causing STEC serotype, E. coli O157:H7 informs most of what is known about STEC . The serotype E. coli O157:H7 was first identified in 1982 and was well studied during that decade . The pathogen rose to infamy in 1993 when a large outbreak occurred across multiple locations of the fast food chain Jack in the Box . The consumption of the chain’s undercooked hamburgers led to illness in more than 600 people . Because of this incident, the way food safety processes are handled, especially the inspection of meat and poultry, have drastically changed . This incident is also the reason why O157:H7 has been so well-studied compared to other STEC serotypes. Among other STEC serotypes E. coli O26 is less likely to cause HUS compared to O157, even though its toxins are similar . Hemolytic uremic syndrome, or HUS, is a severe condition that damages the blood vessels of the kidneys and leads to renal failure.

Once in the body, Shiga toxin can bind to globotriaosylceramide in vascular endothelial cells, and damages those cells by inhibiting protein synthesis. If those cells are part of the kidney, it can lead to HUS . In general, E. coli O157 is more likely to cause severe symptoms than other types of STEC . STEC and may also be of concern in low-moisture foods. While the main reservoir for E. coli O157:H7 is cattle, the pathogen can easily spread through fecal contamination of water and other foods . According to the World Health Organization , this contamination can occur at many stages of growing and processing produce, which has led to increases in outbreaks of the pathogen in fruits and vegetables . Because of the recent outbreaks associated with STEC in low-moisture foods, and the various stages at which contamination can occur, it is important to explore its ability to survive in dried fruit, which can have many processing steps . The detection of STEC can also be culturable or molecular based. Selective media often used for STEC plating include MacConkey agar, violet red bile agar, and Levine’s Eosine methylene blue agar . To differentiate E. coli O157 from other E. coli, sorbitol can be added to the agar since O157 will not usually ferment sorbitol . Because the number of E. coli cells present in food is low, enrichment is very important to make sure that any cells present are detected. Common enrichments forSTEC include brain heart infusion broth, tryptic soy broth, and modified buffered peptone water with pyruvate . For identification in pure cultures, agglutination assays are useful for serotyping . The enzyme-linked immunosorbent assay is becoming more common for identifying STEC. Use of this assay has led to a better understanding of the most common serotypes of STEC. While O157:H7 is the most common STEC serotype associated with disease, there is a decrease in proportion of that serotype when using ELISA compared to culture-based methods . When screening with biochemical tests, most pathogenic E. coli will have negative test results for H2S, urease, arabinose non-fermenting, and indole .

To further determine if pathogenic E. coli is STEC specifically, real-time PCR can be used. The genes that should be targeted during PCR are stx1, stx2, and uidA, with the latter being highly conserved in O157:H7 strains . As mentioned with Salmonella, the main outbreak identification tool used by PulseNet is WGS. It has more differentiation capability than past methods used like PFGE. Abdelhamid et al. looked at a recent outbreak of E. coli O157:H7 from cattle to human and found that WGS was able to distinguish which isolates from the cattle matched the isolates in the infected patients, while the use of PFGE was unable to differentiate between all the isolates tested. Listeria monocytogenes. L. monocytogenes is a gram-positive, non-spore forming bacteria belonging to the family Listeriaceae. It can grow in a temperature range of from -0.4 to 45 °C with optimal growth from 30 to 37 °C and can grow within a pH range of 4.4-9.6, but has optimal growth at 6-8 . L. monocytogenes can grow in foods with a water activity of 0.9 or higher .L. monocytogenes is found in a variety of places, including plants, animals, soil, water, and humans . It can cause listeriosis, which can be a very serious infection in high-risk groups but is unlikely to manifest severely in other groups of people . Foodborne Listeria needs only a few cells to infect and once in the digestive tract Listeria can invade cells and use cell-to-cell transmission to spread to the rest of the body. Symptoms of listeriosis in high-risk individuals can include miscarriage, sepsis, and meningitis, grow bag for blueberry plants while in the rest of the population people may experience only mild gastroenteritis. There is some debate of whether L. monocytogenes poses a significant risk in low moisture foods. There have been no documented outbreaks of L. monocytogenes associated with low-moisture foods and the current prevalence of the pathogen in low-moisture foods is likely low . However, L. monocytogenes can survive for long periods of time in low-moisture foods and there have been recalls associated with L. monocytogenes in these foods, including in dried fruits, nuts, biscuits, and oats . L. monocytogenes is notorious for its ability to grow in cold environments. This is why outbreaks of this pathogen are often found in refrigerated, ready-to-eat foods , as they do not require heating before consumption.

Dried fruits are an RTE and are often stored at refrigerated temperatures by processors, but due to the inability of pathogens to grow at low water activities, L. monocytogenes growth should not be a concern in dried fruits. Pathogen survival is still a concern though, as L. monocytogenes has been shown to have a desiccation tolerance of up to 1 year in certain low moisture foods. For instance, Kimber et al. found that 6 log CFU/gof L. monocytogenes inoculated onto raw almonds did not decline significantly when the almonds were stored at 4 °C for 12 months. Agar used for selective plating of Listeria include Oxford, Modified Oxford, PALCAM, Chromogenic Listeria agar, and lithium chloride-phenylethanol-moxalactam . Selective enrichment can be done with buffered Listeria enrichment broth . Proper subtyping is particularly important in identifying Listeria, as many different strains can have similar phenotypic qualities . The most common serotypes of L. monocytogenes isolated from patients are type 1 and type 4 . There can also be strains that have qualities that are unusual to Listeria that make identification more difficult. The FDA BAM mentions as examples isolates of Listeria innocua that are hemolytic and L. monocytogenes and Listeria welshimeri isolates that are rhamnose negative . When trying to differentiate L. monocytogenes specifically, the species should usually test negative for mannitol and xylose, and should test positive for rhamnose, virulence, and beta hemolysis . Again, sequencing plays an important role in identification of many pathogens such as Listeria. In fact, Listeria was the first bacteria the CDC began using WGS with and has since then spread its use to other organisms including Salmonella and E. coli . Intrinsic factors influencing pathogen survival. Pathogen survival can be influenced by many factors, including aw and pH. In general, the ability of microorganisms to survive common food processes increase when aw is lowered. However, while higher aw promotes growth, high aw also enhances lethality of thermal treatments . The mechanisms for thermal resistance are not completely agreed upon but are shown to be strongly influenced by aw . The lower the aw, the more difficult it is for the number of cells present to decline . For example, Keller et al. found that Salmonella inoculated onto pumpkin seeds became increasingly resistant to thermal inactivation when the aw decreased from its original value of 0.97 to below 0.20. The pumpkin seeds began with a Salmonella population of 7.48 ± 0.57 log CFU/g and dropped to 0.68 ± 0.81 log CFU/g after 6 h or drying at 60 °C . After 6 h, the aw dropped to below 0.20 and no more significant decrease in the Salmonella population was seen during 12 more h of drying at 60 °C . Just knowing the aw alone is not enough information to understand pathogen survival, as water activity is often working in conjunction with other factors such as temperature . pH is also known to have some effect on bacterial survival. While bacteria have a specific pH range in which they can grow, they can survive outside that pH range. Thermal resistance is decreased at lower pH, so pathogens are generally easier to inactivate in more acidic food matrices . Deng et al. inoculated dry infant cereals of pH 4.0 and 6.8 with 6 log CFU/g of E. coli O157:H7. After 24 weeks of storage at 5 °C the cereal with a pH of 4.0 had 3.19 log CFU/g of E. coli, while no E. coli was detected in the cereal at pH 6.8 . The phytochemicals found in dried fruits, including alkaloids, flavonoids, and phenolic compounds, can exhibit antibacterial activity . Jagathambal et al. screened various phytochemicals from dried figs to see if they had any inhibitory effects on various bacteria. The phytochemicals extracted from dried figswere able to inhibit Salmonella spp., Klebsiella spp., Haemophilus spp., and Serratia spp. with a minimum inhibitory concentration of 1.0 mg/mL . Mainasara et al. screened phytochemical from dates to see how inhibitory they could be against pathogens.