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. 2022 Feb 7;60(4):1274–1283. doi: 10.1007/s13197-022-05381-9

Foodborne illnesses of Escherichia coli O157origin and its control measures

Songeeta Singha 1, Rajendran Thomas 1,✉, Jai Narain Viswakarma 2, Vivek Kumar Gupta 1
PMCID: PMC10020406  PMID: 36936116

Abstract

Foodborne illnesses are leading source of morbidity and mortality in both developed and developing nations. Escherichia coli O157 is one of the most reported foodborne pathogen that emerged in the past few decades. South East Asia region suffers the highest average burden of diarrhoeal mortality, especially when it comes to child mortality.Query Many studies were undertaken in the developed nations to evaluate the role of E. coli O157 as one of the etiological agent in foodborne outbreaks. In this article, we discuss the distribution of E. coli O157 serotype in the food chains of South East Asian countries, with a special focus on India where more than half a million child diarrhoeal deaths occurs every year and the reasons for which is often not ascertained to the fullest extent. The article also describes in detail about the various detection methods and control measures with respect to E. coli O157. The aim of this study is to document and highlight the extent of Foodborne infections of E. coli O157 origin and thereby taking effective and proactive preventive measures.

Keywords: Foodborne illness, E. coli O157, Diarrhoea, Contamination, Detection, Control

Introduction

Foodborne illnesses are a major global issue, both in terms of human suffering and economic costs. It is a leading source of morbidity and mortality in both developed and developing nations (Glavin 2003). In the coming years more foodborne pathogens can be expected due to changed production methods and agricultural practices. The globalization of food trade raises the possibility for foodborne hazards to spread throughout the world. Shiga toxin producing Escherichia coli (STEC) serotype E. coli O157 is one of the most reported foodborne pathogen that emerged in the past few decades, causing large scale epidemics and thousands of sporadic cases of diarrhoeal illness every year. E. coli O157: H7 is one amongst the frequently isolated variants which is well studied and characterized serologically by the presence of the somatic O157 and the flagellar H7 antigen. According to the reports, most of the outbreaks were caused by E. coli O157: H7. In the recent years, the United States has reported an E. coli O157: H7 outbreak associated to romaine lettuce, with 60% of the patients being hospitalized (Hoff 2021). The Centers for Disease Control and Prevention (CDC) reported three E. coli O157: H7 outbreaks in 2021, all of which were connected to packed salads, baby spinach, and unknown food sources. However, the pathogenic potential of several other variants that do not react with any of the 53 flagellar antigen groups described for E. coli are also well studied due to their involvement in many E. coli O157 outbreaks. These variants are designated as E. coli O157: NM (non-motile) or E. coli O157: H− has been isolated from fecal samples from hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS) patients in Germany (Feng et al. 1996). As infection with E. coli O157: H7 or E. coli O157: NM mostly depends on the presence of shiga toxin, both variants are normally included under the same serotype and referred herein as E. coli O157. Most of the E. coli O157 related outbreaks have been associated with eating undercooked food (Rangel et al. 2005). Prevalence of this serotype is also noted in animal faeces (Rajkhowa and Sarma 2014). In this article, we discuss the foodborne diseases of South East Asian countries, with a special focus on India. This review also highlights the significance of E. coli O157 in development of diseases in human, its prevalence in the food chain, methods of detection and preventive measures.

Current status of foodborne diseases

The impact of foodborne diseases (FBD) may be assessed in a variety of ways, including yearly instances of illness, severity and mortality. A common measure for assessing disease burden is the Disability Adjusted Life Year (DALY). One DALY is the equivalent of one year of healthy life lost, taking into account the costs of disease and death. In 2006, Foodborne Disease Burden Epidemiology Reference Group (FERG) was established by World Health Organization (WHO) to assess the worldwide burden of FBD. As reported, there are 31 foodborne hazards worldwide, including 11 diarrhoeal disease agents, 7 invasive disease agents, 10 helminthes, and 3 chemicals and poisons (WHO 2019). Among which some of the diarrhoeal foodborne pathogen especially Campylobacter spp., Vibrio cholera, norovirus, EPEC, ETEC and Shigella spp. were each responsible to cause around 3 million DALYs. Salmonella Typhi, Taenia solium, hepatitis A virus, and Paragonimus spp. were among the other foodborne pathogens that contributed significantly to the global burden (Havelaar et al. 2015). Intensive study in the field of foodborne diseases over the last several years has led to broad identification of STEC as a major cause of foodborne diarrhoea. STEC, which includes O157 and several non O157 serotypes, is believed to cause 2,801,000 acute infections, 3890 cases of HUS, 270 cases of persistent ESRD (end stage renal disease), and 230 fatalities each year throughout the world (Majowicz et al. 2014). According to findings based on a study of dysentery cases in India, 37.21% STEC were isolated from patients with abdominal pain, severe bloody diarrhoea, absent or low-grade fever and pus development in stool specimens (Lanjewar et al. 2010). STEC has a higher influence on children than on people of other ages, according to the study. According to the FERG research, children under the age of five are the most affected, accounting for 40% of the foodborne disease burden. It is worth noting that the worldwide mortality rate for people infected with STEC serotype E. coli O157 is greater than those infected with E. coli non-O157 (Majowicz et al. 2014).

Based on the mortality rates of child and adults, all WHO member countries are split into sub-regions. FBD burdens vary substantially between sub-regions with African regions carrying the most loads, followed by Southeast Asia region and the Eastern Mediterranean region (Havelaar et al. 2015). India is one of 11 countries in the SEAR D region, which implies it has a high incidence of child and adult mortality. SEAR D also has the highest average burden of diarrhoeal mortality when it comes to child mortality. Among these, India alone is responsible for more than half a million diarrhoeal deaths (Fig. 1). Kristkova et al. (2017) estimated that there are roughly 100 million instances of FBD in India annually. This corresponds to around one out of every twelve persons being infected with FBD. There are around 120,000 fatalities each year and FBD imposes a burden of nearly 8 million DALYs (Table 1). According to the reports, the number of FBD diseases in 2030 is expected to increase by up to 70% due to population growth and changing consumption habits, resulting in one out of every nine individuals being ill.

Fig. 1.

Fig. 1

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List of top 15 nations based on diarrhoeal child mortality

Table 1.

Annual burden of foodborne disease in India (Kristkova et al. 2017)

Cases Total (in millions) Above 5 yr (in millions) Under 5 yr (in millions)
Foodborne disease 97 58 40
Foodborne diarrhoeal disease 79 44 35
Deaths (foodborne disease) 0.12 0.09 0.03
Deaths (foodborne diarrhoeal disease) 0.06 0.04 0.02
DALYs (foodborne disease) 8.4 5.4 3
DALYs (foodborne diarrhoeal disease) 4.4 2.5 1.9
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DALY Disability adjusted life year

Virulence and pathogenesis

E. coli O157 is well known for its extensive distribution, unusual resistance to various physical and chemical treatments, severity of disease and low dosage infectiveness. Although the overall number of E. coli O157 cases is lesser than that of other enteric pathogens such as Campylobacter spp., Salmonella spp., the diseases caused by E. coli O157 have considerably greater hospitalization and mortality rates (Lim et al. 2010). E. coli O157 infection tends to occur mostly in the summer and autumn seasons. Most instances begin with non-bloody diarrhoea and resolve without further complications. Patients also experience abdominal pain that is more severe than in other types of bacterial gastroenteritis. The time it takes for diarrhoea to appear after consuming contaminated food varies between 2 and 12 d. There are 2 kinds of HUS in children, according to the authors. HUS characterized by diarrhoea, microangiopathic hemolytic anemia, abdominal pain, thrombocytopenia and acute renal failure is described as “typical HUS” or “diarrhoea associated HUS”. On the contrary, “atypical HUS” or diarrhoea-negative HUS refers to children who have symptoms that are identical to that of typical HUS but do not have the symptoms of diarrhoea (Waters et al. 2007). Diarrhoea-associated HUS accounts for 90–95% of children HUS, while diarrhoea-negative HUS accounts for 5–10% (Saxena et al. 2015). The first anomaly in most HUS patients is thrombocytopenia, which is defined by low packed-cell volume with indications of erythrocyte lysis, a low platelet count and elevated serum creatinine (Tarr et al. 2005).Other consequences of shiga toxin such as irritability, lethargy, and dementia, have been anticipated to be induced by fatigue, cerebral microvascularthrombi, ischaemia-hypoxia or the direct neuronal effects of toxin. About 10% of HUS patients experience seizures or coma. During the E. coli O157 outbreak, cardiac dysfunction was observed in around 10% of children with HUS (Tarr et al. 2005).

A Shiga toxin (Stx) mediated endothelial cell damage at the site of infection is the characteristics of E. coli O157 infection. Stx is the main virulence factor of E. coli O157. Stx is one of the potent biological cytotoxin and it is bacteriophage encoded. The acquisition of bacteriophage was a key factor in the emergence of EHEC from EPEC (Lim et al. 2010). Stx subgroups Stx1 and Stx2 share approximately 55% amino acid homology but do not generate cross reactive antibody. While Stx1 is highly conserved, Stx2 has 11 different variations, the most prevalent of which are Stx2c, Stx2d, Stx2e, and Stx2f (Muniesa et al. 2006). Stx is made up of 5 identical B subunits that bind the toxin to the glycolipid receptor globotriaosylceramide (Gb3) on the target cell surface, as well as a single A subunit that cleaves ribosomal RNA to halt protein synthesis in the host cell (Kaper et al. 2004). Portion A is internalized in the cell and transported to kidney, where it damages renal endothelial cells and obstructs the microvasculature. In addition, Stx causes apoptosis in intestinal epithelial cells which is regulated by Bcl-2 family proteins (Kaper et al. 2004). Studies have demonstrated the correlation between human intestinal microvascular endothelial cells (HIMEC) with Stx. It was reported that Stx1 had a 50-fold higher binding affinity for HIMEC than Stx2; nevertheless Stx2 was more toxic to HIMEC than an equivalent quantity of Stx1 (Jacewicz et al. 1999).

However, there are over 200 STEC that have the stx gene but are not linked with human infection, indicating the need for other virulence factors. E. coli O157 adheres to the epithelial cell membrane and induces a condition which is characterized by loss of host cell microvilli and the lesions formed are referred as attaching and effacing (A/E) lesions. According to study, this condition involves Locus of Enterocyte Effacement (LEE). In the intestinal cell, the LEE encoded outer membrane protein (intimin) induces intracellular signalling and triggers cytoskeletal changes (Muniesa et al. 2006; Saxena et al. 2015). LEE encoded intimin, Tir and plasmid of O157 (pO157) were essential to the process of attachment. As per findings, E. coli O157 mutants missing one of these factors could not persist longer in the intestine. In contrast, colonization pattern of E. coli O157 mutants lacking stx and hemolysin (hlyA) were similar to that of wild type (Sheng et al. 2006).

Another virulence factors is a 92 kb plasmid (pO157). Bacterial adhesion to host cells, colonization in intestine and acid resistance is all influenced by genes encoded on the pO157 (Lim et al. 2010). The complete sequence of pO157 revealed 43 open reading frames (ORFs) that showed similarities with known proteins. Only 19 genes, including hemolysin (ehxA), serine protease (espP), zinc metalloprotease (stcE), type II secretion system apparatus (etp), putative adhesion (toxB), catalase peroxidase (katP) and an eae conserved fragment (ecf) were identified as being involved in the pathogenesis of E. coli O157 infections. Incidence of high resistance to multiple antibiotics such as sulfamethoxazole, tetracycline, cephalothin and ampicillin in E. coli O157 isolates shows acquisition of antibiotic resistance genes through pO157. Also, E. coli O157: H7 isolates from sheep and cattle were found to carry the pO157-mediated colistine resistance genes viz. mcr-1, mcr-2, mcr-3, mcr-4, and mcr-5, according to a research by Ayaz et al. (2019).

Routes of human infection

Since its first outbreak in Oregon and Michigan in 1982, beef continues to be the most common vehicle among E. coli O157 outbreak. The reviewed case study of E. coli O157 outbreak in USA from 1982 to 2002 indicated that a total of 350 outbreaks have occurred out of which 71% occurred during summer signifying the effect of environmental conditions upon the growth of this organism (Rangel et al. 2005).Cattle are the primary reservoirs for this organism and the presence of this organism in dairy milk poses risk to infants and children. Source of contamination extends up to wide range of foods, not only beef but other meat such as pork, lamb, chicken etc., vegetables and sprouts. Reported cases of pork that were associated with E. coli O157 infection indicated pork as a potential source or source of cross-contamination for E. coli O157 (Honish et al. 2017). According to reports, vegetables and other food products including lettuce, sprouts, cabbage, radish sprouts, apple cider and apple juice are the common cause of outbreaks of E. coli O157 (Rangel et al. 2005).Contaminated irrigation water and manure in the field, contaminated machinery during processing, wash water, poor handling during transport or contaminated storage facilities are all possible sources of contamination for these products. Kitchen level cross contamination aids in the process of transmission through foods (Rangel et al. 2005).

Despite the fact that the Indian subcontinent has the highest rate of diarrhoea related mortality per year, data on E. coli O157 mortality in India is scarce. In India, STEC was initially isolated in 1999 from animal sources (Gupta et al. 1992).Since then, the serotype O157 has been found to be prevalent in packaged food, meat and meat products, seafood, vegetables, water and fecal samples of animal (Table 2).In an epidemiological survey reported by Sehgal et al. (2008),it was mentioned that E. coli O157 was widely distributed among human and animals, foods and water bodies of India. But a significantly high percentage of E. coli O157 was isolated from meat and meat products, milk and milk products, seafood and water. It is worth noting that E. coli O157 specifically follows an oral route of infection and origin of majority of the E. coli O157 outbreaks were Foodborne. Presence of this serotype in cattle is a source of infection through milk as most of the population in India prefers milk and milk products (Sehgal et al. 2008). This serotype can also be transmitted into meat from the food animal by faeces during slaughtering and processing. The presence of E. coli O157 in a faecal sample followed by the same in a carcass swab clearly indicates unsanitary slaughterhouse operations, implying that inadequate hygiene is the primary cause of microbial contamination in meat. In faecal samples tested from both healthy and diarrhoeic piglets belonging to India’s major pig producing states, 782 isolates of E. coli were obtained and out of which 14.4% had at least one gene characteristic for STEC. The results also reported prevalence of 0.5% E. coli O157 in the total samples (Rajkhowa and Sarma 2014). When a pig becomes infected, it can shed E. coli O157 for up to 2 months, which could lead to horizontal transmission between a naive and infected pig via the formation of contaminated aerosols. Contaminated aerosols are a means of indirect E. coli O157 transmission among pigs (Cornick and VuKhac 2008). It is also important to note that the presence of a few infected animals with high shedding rates is far more important than the presence of other shedding animals. This serotype was isolated from faecal samples of sheep and goat in India as well (Table 2).Though sheep and goats are also considered as asymptomatic carriers for E. coli O157, according to an experiment conducted in India, the percentage of sheep that shed E. coli O157 in their faeces was greater (3.67%) than that of goats (only 2.88%).

Table 2.

Prevalence of E. coli O157 reported in India

Details of the source Region/states Samples /E. coli isolates E. coli O157 isolates (%) References
Meat (beef and chevon) Puducherry 115 13.91 Vijayan et al. (2017)
Packaged food (frozen, cooked, semi cooked, uncooked and ready to eat) Assam 83 15.66 Sharma et al. (2019)
Ganges river Varanasi, UP 5 100 Hamner et al. (2007)
Faecal sample (sheep and goat) North Eastern Karnataka 967 3.3 Vinay et al. (2018)
Cabbage, radish, lettuce, broccoli and cucumber Coimbatore, Tamil Nadu 603 6.1 Sai (2019)
Meat Mangalore 103 0.97 Dhanashree and Mallya (2008)
Faecal sample (pig) Assam, Arunachal Pradesh, Meghalaya, Nagaland 782 0.5 Rajkhowa and Sarma (2014)
Meat Across the country 1376 0.9 Sehgal et al. (2008)
Milk and milk products Across the country 553 1.8 Sehgal et al. (2008)
Seafood Across the country 190 8.4 Sehgal et al. (2008)
Water Across the country 486 1.6 Sehgal et al. (2008)
Raw vegetables Assam 350 2.57 Nath et al. (2019)
Salad vegetables Dhanbad, Bihar 480 0.62 Mritunjay and Kumar (2017)
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Previous studies indicated some possible transmission route which includes direct contact with the animal or person-to-person contact, contaminated water and manure etc. E. coli O157 was reported to exist in water for long periods of time while retaining their pathogenic potential. A protective capacity of biofilms produced by E. coli allowed them to persist in water distribution systems (Daly et al. 1998). As water bodies serve as environmental reservoirs of E. coli O157, it can easily contribute to subsequent infection of livestock with E. coli O157; alternatively animals may contaminate these water bodies with fecal material, feed, bedding or saliva. Contaminated water emphasizes the importance of control of E. coli O157 in source of water supply to the farm or drinking water.

In India with total population of 1.34 billion people, more than 58% population do not have access to safe drinking water with microbiological counts within prescribed values in municipal drinking water. Identification of potential pathogenic isolates from extensively used water sources indicates that E. coli O157 might be a significant cause of incidence of waterborne diseases among the residents who live near river bodies but this is yet to be acknowledged as public health concern in India. The existence of E. coli O157 was reported in high numbers throughout the Ganges in Varanasi. The water samples were also tested positive for stx1, stx2, and eae gene (Hamner et al. 2007). Recently Pushpakanth et al. (2019) also reported fresh vegetables of the south India were highly prone to E. coli O157 from soil and irrigated water. They also pointed out the contribution of wash water used for carrot in disseminating the same. Manure harboring E. coli O157 might be a source of infection for vegetables or fruits that are not usually prepared before consumption. E. coli O157 has been seen surviving for a year in manure-treated soil and for 21 months in raw manure that had not been composted. In bovine faeces inoculated initially with 105 CFU/g, E. coli O157 survived for 70, 56 and 49 d at 5, 22 and 37 °C, respectively (Bach et al. 2002). Hence, good agricultural practices and good hygiene post-harvest practices have to be imposed in the vegetable production system to avoid the pathogen entry.

Methods of detection

The common methods which are in use for detection of E. coli O157 are presented in Table 3 and are described in detail below.

Table 3.

List of methods used for the detection of E. coli O157

Method Approx. detection time Limit of detection References
Plating/culturing 5 d Low CFUs Doyle and Schoeni (1984)
MUG dependent fluorogenic assay 20 min–72 h 103–104 cells/g Thompson et al. (1990)
Hydrophobic grid membrane filter-immunoblot 3–4 days 1–10 Cells/g Doyle and Schoeni (1987)
Enzyme-linked immunomagnetic electrochemical (ELIME) detection 80 min 103 cells/ml Gehring et al. (1999)
Fluorescent bacteriophage assay (FBA) 7–12 h 2.2 CFU/g 101–102 CFU/ml Goodridge et al. (1999)
Latex agglutination 24 h 1 CFU March and Ratnam (1989)
Immuno-magnetic separation (IMS) 48 h 10 CFU/ml Fratamico et al. (1992)
BAX PCR based system 2–4 h 1–10 CFU/g Shearer et al. (2001)
MAb-based ELISA 24 h 104–105 cells/ml Padhye and Doyle (1991)
PCR assay 2–4 h 103 CFU Ayaz et al. (2014)
Reverse transcriptase-polymerase chain reaction assay 2–4 h 107 CFU Yaron and Matthews (2002)
Real time PCR 2–4 h 104 cells Spano et al. (2005)
Nucleic acid microarrays 2–4 h 102 CFU/ml Call et al. (2001)
Multiplex PCR-based assay 2–4 h 1–2 CFU/ml Campbell et al. (2001)
Molecular beacons (MBs) based real time PCR assay 2–4 h 1 CFU/ml Fortin et al. (2001)
Matrix-assisted laser desorption-ionization time of flight mass spectrometry (MALDI-TOF MS) 1 h 1 CFU/ml Rychert (2019)
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Conventional methods

Conventional detection method is mainly based on culturing followed by isolation and biochemical identification. The three most important phenotypic characteristics that segregate E. coli O157 from rest of the genus E. coli are absence of β-glucuronidase enzyme, inability to ferment sorbitol and poor or no growth at temperature beyond 44 °C (Doyle and Schoeni 1984). A persistent phenotypic feature of E. coli O157 is the lack of sorbitol fermentation within 24 h and the efficacy of Sorbitol MacConkey agar (SMAC) is useful for selection of Non Sorbitol Fermenting (NSF) E. coli O157. It was also observed that distinctive growth of the E. coli O157 as NSF colonies on SMAC medium, allows presumptive identification of this serotype with a low false-positive rate. Evidence of Sorbitol Fermenting (SF) E. coli O157, on the other hand, may risk the SMAC-based isolation technique. According to Ayaz et al. (2014), the presence of SF E. coli O157: H7 isolates from cattle emphasizes the need for an alternate methods for the shiga toxigenic SF E. coli O157: H7. Moreover, According to some studies the specificity of the SMAC medium for E. coli O157 serotype is only 45–52% due to the presence of NSF organisms other than E. coli O157. E. coli O157 has been reported to be uniformly 4-methylumbelliferyl-β-D-glucuronide (MUG) negative unlike 96% of the E. coli isolates (Thompson et al. 1990). The fluorogenic end product of enzyme β-D-glucuronidase and substrate MUG is detectable under UV light. This characteristic has been used for direct isolation of MUG negative E. coli O157 from food samples. In recent years chromogenic agar has also evolved and showed improved diagnostic efficiency in diagnosing of E. coli O157 infections. Rainbow agar O157, BBL CHROMagar O157, showed a 100% negative predictive value and a higher diagnostic efficiency than SMAC.

Immunological methods

Several immunological approaches have been developed to confirm presumptive E. coli O157 isolates from conventional culturing methods. Direct Immunofluorescence (DIF) aids in the identification process of E. coli O157. In many cases, SMAC agar as a primary isolation medium combined with DIF resulted in fast identification of the E. coli O157 serotype in less than 24 h. Previously described Monoclonal antibody (MAbs) was mostly reactive to the somatic O157 antigen. As a result, they responded to all E. coli strains from serotype O157 and exhibit cross reactivity with group N salmonellae. Padhye and Doyle (1991) constituted a monoclonal antibody (MAb 4E8C12) specific solely to E. coli O157: H7 and O26: H11. The procedure involves immunizing BALB/c mice with a rough strain of E. coli O157: H7 which lacks a part of the lipopolysaccharide (LPS) layer of the outer cell membrane. Specificity of MAb 4E8C12 was determined by ELISA and was indicated by the authors that this antibody could be used for the detection of E. coli O157: H7 and O26: H11 from clinical and food samples.

Isolation of a specific bacterial species from a mixed culture, such as food, is usually ineffective without a pre-enrichment step. When magnetic separation is used instead of a pre-enrichment stage, organisms can be targeted straight from the sample or pre-enrichment medium. Immuno Magnetic Separation (IMS) is the potential method to capture and detect dead or severely damaged cells that the conventional detection method cannot detect. It is reported that the ratio of IMS particles to target cells should be 3:1 to 20:1. According to the study, increasing the incubation time significantly improves target cell recovery. However, when there is a high concentration of other competing flora in the matrix, such as in meat, it is advisable to choose a shorter incubation time to minimize the level of nonspecific binding (Fratamico et al. 1992). Screening method combining a commercially available reactive disc blot ELISA (3 M Petrifilm™ Test Kit) for E. coli O157 antigen with a conventional method was also reported for the isolation of E. coli O157. A commercial latex agglutination test kit was evaluated based on its sensitivity as well as specificity against clinical isolates of E. coli O157. The specificity was determined using the isolates from HC outbreak in which it correctly identified and tested positive to all NSF colonies produced by E. coli O157 and gave a clear cut negative result to NSF colonies from non E. coli O157 (March and Ratnam 1989).

Several approaches have been introduced combining selectivity of immunoassay with the rapidity and sensitivity of electrochemistry to detect target organism. Gehring et al. (1999) applied Enzyme-linked Immunomagnetic Electrochemistry (ELIME) for the rapid detection of E. coli O157 cells in buffer or porcine carcass wash water. The ELIME methodology involved ‘sandwiching’ of E. coli O157 cells between immunomagnetic beads and an alkaline phosphatase (AP)-conjugated antibody.The custom-synthesized AP substrate, p-aminophenyl phosphate (pAPP) has been used for the ELIME detection of S. typhimurium. However, for the detection of E. coli O157 use of pAPP was effectively replaced by commercially available 1-Naphthyl phosphate. Similarly, Fluorescent Bacteriophage Assay (FBA) is another example of combination method that involves immunomagnetic separation and highly specific fluorescently stained bacteriophage (Goodridge et al. 1999).

PCR based methods

Bacterial genotyping eliminates many of the disadvantages of phenotypic assays, such as differences in enzyme activity depending on the medium used, the emergence of biochemical mutants and the occurrence of closely related strains with the same phenotypic features but differing in genotype. Several genes and molecular markers have been targeted in order to develop molecular techniques for identifying E. coli O157. In a study, Ayaz et al. (2014) evaluated the sensitivity of a PCR procedure for detection of E. coli O157 in cattle and slaughterhouse wastewater. As described by the authors, this PCR approach correctly assessed the presence of genes responsible for identification, virulence factors, intimin variations, and shiga toxin variants in the isolates. Ability of a PCR detection method is completely dependent on the extraction procedure of genomic material. The ideal DNA extraction procedure yields the most amplifiable DNA with minimal co-extraction of potential PCR inhibitors that could reduce the sensitivity of the PCR method. In their study, Holland et al. (2000) compared commercial kits and one in-house technique for evaluating yield of amplifiable DNA for PCR. The in-house method and all commercial kits shared comparable specificities, which were in the range of 98–100%. With the exception of one technique; all methods had similar sensitivities within the range of 83–86%. It is also reported that PCR results are also dependant on source of the isolates and the sample preparation techniques. According to the study, the extraction procedures such as centrifugation and IMS were successful in minimizing false positive results following the addition of washing stages before DNA extraction (Fratamico et al. 1992).

Several variations of the conventional PCR have recently evolved; allowing for more sensitive detection techniques to be developed. The most prevalent methods are multiplex PCR and real-time PCR. The former permits numerous targets to be co-amplified in a single PCR by combining or multiplexing primer pairs. Hu et al. (1999) have developed a PCR by using primer sets that amplified the genes viz. eaeA, slt1, slt2, fli C and rfb E. These sets of primers were evaluated analyzing both E. coli O157 and non E. coli O157 isolates from bovine faeces in which this method was specific to E. coli O157 and its virulence factors and was able to distinguish O157 from other serotypes. In a study, efficacy of multiplex PCR in combination with enrichment step was evaluated in E. coli O157 spiked environmental samples. According to the authors, amplification of the target gene was dependent on initial cell concentration, but environmental stressors such as starvation had no effect on this approach of detecting the serotype (Campbell et al. 2001). In a method developed by Yaron and Matthews (2002), the effects of temperature and nutrient limitation on the identification of viable cells were investigated using reverse transcriptase PCR. The rfbE gene was shown to be the most appropriate target for detecting viable E. coli O157 under the circumstances studied. As far as detection was concerned, realtime PCR assay was found effective to detect E. coli O157 in wastewater at a contamination level of 104 CFU/ml (Spano et al. 2005). The recent discovery of fluorogenic nucleic acid probes such as molecular beacons (MBs) adds a new dimension to the PCR. MBs combined with real time PCR were assessed in the detection of E. coli O157 in milk and juice samples. The highly specific nature of MBs has the capability to discriminate even a single base mismatch in the target sequence (Fortin et al. 2001). BAX is one of the commercially available PCR based systems developed for the rapid detection of bacteria in raw and finished products as well as environmental samples. While evaluating the presence of pathogenic strains in fresh vegetables and fruits, Shearer et al. (2001) stated that BAX was more sensitive than the culture-based method in detecting E. coli O157 in green pepper, carrot, radish, and sprout samples.

Control measures

Most of the Foodborne diseases are sporadic and often under reported in India; however a nationwide surveillance study reported 13.2% foodborne illnesses at the household level (Kohli and Garg 2015). The Food Safety and Standards Act was the first comprehensive approach to tackle the widespread incidence of FBD and large scale prevalence of dietary deficiencies in India. As per the Food Safety and Standards Authority of India (FSSAI) standard, E. coli O157 should be absent in 25 g of meat and meat products. The act also intends to meet the standards of food trade within the country. Key strategies to ensure food safety also include an effective surveillance system linked to a food safety database information system. In order to ensure the microbiological safety of food, it is essential to investigate the prevalence of the causative agent in food. Implementation of Hazard Analysis and Critical Control Point (HACCP) in food industry is mandatory to identify potential food safety hazards, measures for their control and determine critical control points. HACCP approach has been adopted by Codex, USPA, EU and other national and international regulatory and advisory bodies as the cornerstone of microbiological food safety. Risk assessment, however, is a more recent, scientific development in hazard identification. HACCP and risk assessment are complementary to each other but differs fundamentally, with risk assessment focusing on single pathogen and HACCP taking account of multiple hazards for a single product in a particular manufacturing facility (McClure et al. 2012).

In order to control the cross contamination, control measures need to be taken. Water sources contaminated with E. coli O157 should be avoided for irrigation of food crops. Since fresh vegetables and fruits are consumed raw, thorough washing of these produce with saltwater is more efficient than washing with tap water. Furthermore, it was found in a study that washing vegetables with an acidified chlorite solution decreased the population of E. coli O157 by 3.0 log CFU/g. Washing treatments including chlorine, electrolyzed water and chlorine dioxide solutions were shown to be efficient in decreasing the quantity of E. coli O157 in lettuce leaves (Keskinen et al. 2009). However, in some instances the treatment efficiently reduces the serotype but softens the leaves. Alternative methods were also reported to control the E. coli O157: H7 counts in foods without affecting the sensory characteristics. Bacteriophages have been widely used as a biocontrol way to decrease E. coli O157: H7 counts without affecting the sensory properties of food. The use of a virulent phage against E. coli O157: H7 was reported in Turkish raw meatballs, where higher phage concentration resulted in a considerable reduction in E. coli O157: H7 levels (Gencay et al. 2016). It is claimed that phages isolated from poultry slaughterhouse wastewaters are effective against Listeria monocytogenes in chicken carcasses (Cufaoglu and Ayaz 2019). Furthermore, oscillation method, gamma irradiation, and high pressure processing are some of the often used techniques to eliminate E. coli O157 in raw and processed food (Kristkova et al. 2017). In order to eradicate E. coli O157 in fecal excretion of food animal, vaccination of the animal is possibly the most effective and cost-efficient technique. Sharma et al. (2018) formulated a vaccine with killed whole cells of hha mutant E. coli O157 emulsified with an adjuvant. The vaccine was capable of inducing an O157-specific immune response in the host, which either directly or indirectly decreases E. coli O157 shedding in faeces. The efficacy of direct administration of polymyxin B and chlorhexidine to the rectal mucosa to decrease or eradicate E. coli O157 from calves was also evaluated. Both therapeutic drugs were able to reduce shedding of the serotype in faeces by eradicating at the terminal rectum (Naylor et al. 2007).To avoid contamination, it is suggested that whole meats and minced meats should be cooked to a temperature of at least 70 °C throughout the product and this temperature should be maintained for at least 2 min. Hygiene practices in everyday life are also effective in reducing the secondary transmission. According to a disease transmission model, even a moderately effective method to prevent secondary transmission might result in a 5–11% reduction in symptomatic cases. These methods include frequent hand washing, avoiding contact with people who have diarrhoea (of any etiology), preparing hygenic food and isolating those who have any gastrointestinal sign or symptom (Seto et al. 2007).

Future scope

Given the prevalence of E. coli O157 in the food chain and water sources, corrective measures and effective monitoring system to resolve the problem is required. Incidence of E. coli O157 fatalities may be occasional as reported but chances of an outbreak in near future cannot be denied. Attention should be paid in developing relevant institutional and human resources needed to monitor the serotype in food and clinical samples effectively. With the help of advance and cost effective techniques for detection, screening of E. coli O157 would be possible in large number, especially in resource limited developing countries.

Author contributions

All the listed authors have equally contributed.

Funding

The required fund has been received from ICAR-National Research Centre on Pig.

Availability of data and material

Available with the institute.

Declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Consent for publication

Approved by the Performance Monitoring and Evaluation (PME) cell of the institute for sending for publication to JFST.

Ethical approval

All the research works carried out at Food Quality Control Lab is approved by Institute Animal Ethics Committee and Institute Biosecurity Council.

Footnotes

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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