**5. Detection of** *E. coli* **O157:H7**

#### **5.1 Cultured-based detection**

O157 STEC can usually be differentiated from most commensal *E. coli* by their ability to ferment sorbitol when plated on a sorbitol-containing agar.

For isolation this strain, samples are plated onto a selective and differential media, such as sorbitol-MacConkey agar (SMAC), cefixime tellurite-sorbitol MacConkey agar (CT-SMAC), CHROMagar O157, or Rainbow agar. After incubation for 16–24 hours at 37°C, the plate is being examined for possible O157 colonies, which are colorless on SMAC or CT-SMAC and are mauve or pink on CHROMagar O157 [26].

Non-motile flagella-less (H-) sorbitol-fermenting STEC O157 might not grow on CT-SMAC agar because of their susceptibility to tellurite [27].

In the laboratory, culture and biochemical analysis is considered as the "gold standard" for the identification of STEC. Selective media, such as SMAC and CT-SMACK can be used to identified O157:H7 STEC because of the ability to ferment sorbitol within 24 hours [28].

Guidance for public health laboratories on the isolation and characterization of Shiga toxin-producing *Escherichia coli* (STEC) from clinical specimens are given in **Table 1** [29].

#### **5.2 Nucleic-acid-based detection**

Recently, PCR methods (like real-time PCR and conventional PCR) have been developed to test the samples for the presence of Stx genes [30].

This method is inexpensive and easy to perform. During the protocol multiple primer sets in a single PCR reaction in order to detect different types and subtypes of Stx genes in a certain sample. But we have to keep in mind, the detection of Stx mRNA is not possible because they have not been expressed yet and there's a possibility of having a false negative test. Also analysts must be aware of the presence of cryptic bacteriophages which are prophages that have become trapped within a bacterial genome [31].


#### **Table 1.**

*Guidance for public health laboratories on the isolation and characterization of Shiga toxin-producing*  Escherichia coli *(STEC) from clinical specimens [29].*

#### **5.3 Detection by monoclonal antibodies**

Detection of Shiga-toxin in clinical samples has been approved by the FDA [27]. These kits can detect Shiga-toxin in the enrichment samples, although none of

them can distinguish the seven subtypes of Stx2 or the three subtypes of Stx. In 2015, researchers designed sandwich ELISAs capable of detecting and distinguishing between stx2 subtypes a, c, and d [32].

These antibodies provide a significant way to test the samples as fast as possible, even including samples from beef and pure culture [33].

#### **5.4 O- and H-antigen determination**

The most common method used in clinical laboratories when samples suspected to O157:H7 are being tested, is the O-antigen determination which is run by latex agglutination. These latex particles are coated with antibodies against the O157 antigen and when they are mixed with bacterial growth, O157 STEC bacteria will bind to the latex particle to produce visible agglutination which means positive reaction [34].

H7-specific antisera for latex agglutination are available for O157 but unlike the previous method, detection of flagellar antigens may be difficult and usually it is being done for non-O157 outbreaks [34].

## **6. Epidemiology and outbreaks**

STEC infection causes a wide spectrum of illnesses, such as non-bloody diarrhea, hemolytic uremic syndrome (HUS), and hemorrhagic colitis (HC) [35].

Many non-O157:H7 STEC strains may also cause HUS but the majority of diarrheaassociated HUS cases in the US are caused by infection with O157:H7 STEC [36].

Escherichia coli *O157:H7 and Its Effect on Human Health DOI: http://dx.doi.org/10.5772/intechopen.101825*

STEC are found in the intestines of healthy animals and are easily transmitted to humans by consumption of contaminated food or water, or even through direct contact with infected animals or persons [37].

Undercooked beef especially ground beef plays an important role in many O157:H7 STEC outbreaks, although other foods including unpasteurized juice, raw milk, and raw produces (e.g., lettuce) have been implicated in outbreaks too [38–40].

For the years 1998 and 1999 data about in implicated vehicles in outbreaks of *E. coli* O157:H7 exist and are given in **Table 2**.

CDC officials in several states, and the U.S. Food and Drug Administration (FDA) have collected data to investigate a multistate outbreak of *E. coli* infections linked to cake mix. As of July 27, 2021, 16 people infected with the outbreak strain of *E. coli* O121 have been reported from 12 states. Illnesses started on dates ranging from February 26, 2021 to June 21, 2021 and this outbreak is over right now [41].

In December 22, 2020 the FDA and CDC investigated a multistate outbreak of *E. coli* O157:H7 infection linked to leafy greens, a total of 40 people infected with the outbreak strain of *E. coli* O157:H7 were reported from 19 states. Illness started on dates ranging from August 10, 2020 to October 31, 2020. Ill people ranged in age from 1 to 85 years [42].

Outbreaks of *E. coli* infection linked to leafy greens (which often eaten raw with no cooking) including various types of lettuce such as romaine or iceberg lettuce, spinach, and mesclun mix in Canada and United States, are known as critical issues since 2008 [43].

Of the 57 *E. coli* infection outbreaks identified, 48 were attributed to *E. coli* O157 and the most of the causative agents (45 of the 48 outbreaks) were identified as *E. coli* O157:H7 and the other nine outbreaks were attributed to non-O157 *E. coli* [44].


#### **Table 2.**

*Food vehicles implicated in outbreaks of* E. coli *O157:H7, United States, 1998–1999.*

In the United States since 2008, 37 outbreaks of *E. coli* O157:H7 infection linked to leafy greens were identified: 5 linked to iceberg (13.5%), 11 linked to romaine lettuce (29.7%), and 21 linked to other or unspecified types of leafy greens (56.8%). These 37 outbreaks resulted in 1070 illness cases: 491 linked romaine lettuce (45.9%), 144 linked to iceberg lettuce (13.5%), and 435 linked to other or unspecified types of leafy greens (40.7%) [45].

Information on which month the outbreaks occurred is available for 17 of the 18 outbreaks linked to romaine lettuce in Canada and the United States from 2008 to 2018. The majority of these outbreaks happened during two seasons: eight occurred in the spring (March to June) and eight occurred in the fall (September to December) [46].

A possible theory for the distribution of *E. coli* O157:H7 illness cases observed in Canada could be related to the commercial distribution of lettuce. Lettuce imported from U. S. lettuce-growing regions can travel long distances to reach Canada and even distances farther t the eastern part of the country [47].

In the spring 2018 outbreaks in U.S., trace back investigation identified 36 growing fields on 23 farms in the Yuma, AZ, growing region as potential sources of contaminated lettuce. Growers reported the following common elements: romaine lettuce was grown under conventional agricultural practices; Colorado River water via an open irrigation canal was used to irrigate the romaine lettuce and to dilute agricultural chemicals; and overhead sprinkler irrigation was used during the germination of romaine lettuce followed in most fields by furrow irrigation [48].

In November 5, 2019 to November 16, 2019, CDC and FDA investigated a multistate outbreak of Shiga toxin-producing *E. coli* O157:H7 infection started in Georgia, Illinois, Minnesota, North Dakota, and Wisconsin. A total of 10 people ranged from 21 to 91, with a median age of 33, infected with the outbreak strain of *E. coli* O157:H7 were reported and 60% were female. Four of 10 ill people were hospitalized, including one person who developed hemolytic uremic syndrome. No deaths were reported. Information collected during the investigation showed that Fresh Express Sunflower Crisp chopped salad kits were the likely source of this outbreak. Of the 10 ill people with information available, all 10 reported eating any leafy greens in the week before their illness started. Eight ill people specifically reported eating a Fresh Express Sunflower Crisp chopped salad kit.

As of January 15, 2020, this outbreak appears to be over [49].

In December 2020, CDC, U.S. Food and Drug Administration, and public health regulatory officials reported an outbreak of *E. coli* O157:H7 in several states.

Public health investigators used the national PulseNet system to identify illness that may have been included in this outbreak. PulseNet system is the subtyping network of public health and food regulatory agency laboratories conducted by CDC, and with the help of the whole genome sequencing (WGS) method, analyzing the DNA fingerprinting is being done by official investigators. Molecular Investigations showed that people in this outbreak were more likely to share a common source of infection. As of December 16, 2020, a total of 32 people infected with the outbreak strain of *E. coli* O15:H7 were reported from 12 states. Illnesses started on dates ranging from June 6, 2020 to October 25, 2020. Ill people ranged in age from 2 to 75 years, with a median age of 27 years, and 72% were female. Of 29 ill people with information available, 15 were hospitalized and one developed hemolytic uremic syndrome (HUS). One death was reported from Michigan.

The officials interviewed ill people to determine what they ate, they reported variety of food items. Several ill people also reported eating at the same restaurant with common foods. As of December 18, 2020, this outbreak is over and that ended unknown, before enough information was available for investigators [50].

#### Escherichia coli *O157:H7 and Its Effect on Human Health DOI: http://dx.doi.org/10.5772/intechopen.101825*

In November, 2021 CDC and FDA collected data to investigate a multistate outbreak of *E. coli* O157:H7 infections. Epidemiologic and laboratory data show that Josie's Organics prepackaged baby spinach may be contaminated with *E. coli* and may be making people sick. As of the November 15, a total of 10 people infected with the outbreak strain of *E. coli* O157:H7 have been reported from seven states. Illnesses started on dates ranging from October 15 to October 27, 2021. Sick people range in age from 2 to 71 years, with a median age of 26, and 70% are female. Of eight people with information available, two have been hospitalized and no deaths have been reported.

Public health investigators are using the PulseNet system and WGS method to identify illnesses that could be part of the outbreak, which showed that bacteria from sick people samples are closely related genetically, that suggests that people in the outbreak got sick from the same food. This outbreak is not over yet and CDC is advising people not to eat, sell or serve Josie's Organics prepackaged with best by date of October 23, 2021 [51].

On December 11, 2017 the Public Health Agency of Canada (PHAC) announced an outbreak of 21 STEC O157:H7 infections in three provinces linked to romaine lettuce.

This outbreak appears to be over as of January 25, 2018, and 25 people infected with the outbreak strain of STEC O157:H7 were reported [52].

On January 4, 2017 to April 18, 2018 *E. coli* O157:H7 infections linked to I.M. Healthy Brand SoyNut Butter was started and there were 32 people infected in Arizona, California, Florida, Illinois, Oregon, and Virginia and 9 people developed HUS. The source of the infection was a nut-free substitute for peanut butter and this outbreak seems to be over [53].

On June 27, 2016 to September 10, 2016, there were 11 reports based on *E. coli* O157:H7 infections in Connecticut, West Virginia, Pennsylvania, and Massachusetts. Epidemiologic and laboratory evidence indicated that beef products produced by Adams Farm Slaughterhouse in Athol, Massachusetts, were likely source of this outbreak and one ill person developed HUS. On October 19, 2016 officials declared this outbreak over [54].

On October 6, 2015 to November 3, 2015, there were a total of 19 people infected with the outbreak strain of Shiga toxin-producing STEC O157:H7 in Missouri, Colorado, Utah, Virginia, Washington, and Montana, ranged in age from 5 to 84 years, with a median age of 18. Two ill people developed HUS. The epidemiologic evidence collected during this investigation suggested that rotisserie chicken salad made and sold in Costco stores was the likely source of this outbreak. This outbreak seems to be over reported to CDC [55].

On May 19, 2014, a total number of 12 persons were infected with the outbreak strains of Shiga toxin-producing *E. coli* O157:H7 were reported from Michigan, Missouri, Ohio, and Massachusetts. No ill people developed HUS. Federal officials indicated that contaminated ground beef produced by Wolverine Packing Company was the likely source of this outbreak of STEC O157:H7 infections. On June 20, 2014 this outbreak was over reported by CDC [56].

A total of 33 ill persons infected with the outbreak strain of STEC O157:H7 were reported from Arizona, California, Texas, and Washington, on November 10, 2013. Federal officials indicated that consumption of two ready-to-eat salads, Field Fresh Chopped Salad with Grilled Chicken and Mexicali Salad with Chili Lime Chicken, produced by Glass Onion Catering and sold at Trader Joe's grocery store locations, was the source of this outbreak of STEC O157:H7 infections and even two ill people developed HUS. This outbreak appears to be over [57].

From October 18, 2012 to November 12, 2012, a total of 33 people infected with the outbreak strain of STEC O157:H7 were reported from Connecticut, Massachusetts, New York, Pennsylvania, and Virginia and two ill persons developed HUS. Epidemiologic investigation conducted by officials in local linked this outbreak to prepackaged leafy greens, produced by State Garden of Chelsea, Massachusetts. Testing conducted by the New York Department Health Wadsworth Center Laboratories isolated the outbreak strain of STEC O157:H7 from four leftover packages of Wegmans brand Organic Spinach and Spring Mix blend collected from four ill person's homes [58].

As of March 22, 2011, 14 persons infected with the outbreak strain of *E. coli* serotype O157:H7 have been reported from Maryland (three cases), New Jersey (two cases), North Carolina (one case), Ohio (two cases) and Pennsylvania (six cases) and none have reported HUS. Reported dates of illness onset range from January 10, 2011 to February 15, 2011. Collaborative investigative efforts of local, state, and federal officials have associated this outbreak with eating Lebanon bologna, produced by Palmyra Bologna Company, which is a fermented, semi-dry sausage. In an epidemiologic study conducted during March 15–18, a total of 13 ill persons reported about the common disease [59].

From October 16, 2010 through October 27, 2010, 38 persons infected with the outbreak strain of *E. coli* O157:H7 have been reported from New Mexico (3 cases), Arizona (19 cases), California (3 cases), Colorado (11 cases), and Nevada (2 cases) with one case of HUS. The officials linked this outbreak to Bravo Farms Dutch Style Gouda Cheese purchased from a Costco stores [60].

As of Tuesday, June 30, 2009, 72 persons infected with *E. coli* O157:H7 with a particular DNA fingerprint have been reported. The number of ill persons in each state is as follows: Arizona (2 cases), California (3 cases), Georgia (1 cases), Illinois (5 cases), Kentucky (2 cases), Montana (1 cases), Maine (3 cases), Maryland (2 cases), Nevada (2 cases), Ohio (3 cases), Oklahoma (1 cases), Utah (4 cases), Texas (3 cases), New Jersey (1 case), and Wisconsin (1 case). Ill persons range in age from 2 to 65 years, Thirty-four persons have been hospitalized and 10 people have developed HUS. The epidemiological study indicated a strong association with eating raw prepackaged cookie dough. Most patients reported eating refrigerated prepackaged Nestle Toll House cookie dough products raw. This outbreak is over [61].

As of July 17, 2008, 49 confirmed cases have been linked both epidemiologically and molecular fingerprinting to *E. coli* O157:H7. The number of cases in each state is as follows: Georgia (4 cases), Indiana (1 case), Kentucky (1 case), Michigan (20 cases), Ohio (21 cases), and Utah (1 case). Their illnesses began between May 27 and July 1, 2008. Twenty-seven persons have been hospitalized and one person developed HUS. The officials reported the ground beef sold at Kroger Co was the main source of the infection. This outbreak is over [62].

On December 14, 2006, 71 persons with illness associated with the Taco Bell restaurant outbreak have been reported to CDC from five states: New Jersey (33 cases), New York (22 cases), Pennsylvania (13 cases), Delaware (2 cases), and South Carolina (1 case). A total of 71 ill persons, 53 were hospitalized and 8 developed HUS. This outbreak has ended [63].

### **7. Hemolytic uremic syndrome**

Hemolytic uremic syndrome (HUS) is complication of Shiga-toxin producing *E. coli* (STEC) infection. Prompted by the finding of an STEC in the stool of a patient who died from HUS Karmali et al. examined the samples of different cases of HUS and found evidence of STEC infection in 11–15 patients [64].

HUS actually develops 1 week or more after diarrhea begins. Due to the use of the immunomagnetic separation (IMS) the isolation of O157:H7 in the stools from patients with HUS has been increased dramatically [65].

Escherichia coli *O157:H7 and Its Effect on Human Health DOI: http://dx.doi.org/10.5772/intechopen.101825*

You can see the timeframe of the development and evolution of STEC-HUS (**Figure 2**).

The pathogenesis is related to the endothelial cell damage caused by Shiga-toxin which is produced in the gastrointestinal tract. These damaged cells become separated from the basement membranes of the glomeruli and in blood vessels to other organs. Clinically HUS from *E. coli* O157:H7 first appears similar to other common severe gastroenteritis, stools may be bloody, fever is absent, and thrombocytopenia as anemia is a hallmark. Edema is common in later stages and also blood loss via gastrointestinal tract and small blood vessels due to active hemolysis is common too. The *E. coli* O157:H7 can easily be cultured from feces. For more information please see (**Table 3**) which shows the distribution of children and adults in studies of STEC-HUS [67].

#### **Figure 2.**

*Timeframe of the development and evolution of STEC-HUS. The timeframe and proportion represented are based on median values and are highly variable, depending on strain, epidemiological, and individual patient characteristics [66].*


#### **Table 3.**

*Distribution of children and adults in STEC-HUS cases [67].*


#### **Table 4.**

*Annual HUS in U.S. incidence per 1,000,000 children [67].*

HUS primarily affects the kidneys, however it can also lead to sepsis and neurological damages [66]. In (**Table 4**) there are some information based on Annual HUS incidence per 1,000,000 children in U.S. [67]. All the data based on HUS cases in last 10 years, are given in Section 6.
