Pathogens and Toxins in Foods: Challenges and Interventions

Campylobacter species have a chemoorganotrophic metabolism, and energy is derived from amino acids or tricarboxylic acid cycle intermediates due to their inability to oxidize or ferment carbohydrates. The majority of Campylobacter spp. reduce nitrate and nitrite. Campylobacter jejuni contains two flagellin genes, flaA and flaB; the wild-type bacterium expresses flaA only, but flaB can be expressed under certain conditions. The flagellum of Campylobacter spp. plays a much more important function than just motility. C. jejuni flagella may also play a role in the dissemination and internalization of the organism. Toxin production plays a role in pathogenicity. In regard to C. jejuni, the organism synthesizes several toxins, classified mainly as enterotoxins or cytotoxins. The prevalence of Campylobacter contamination of carcasses and poultry products can vary greatly, depending on the sensitivity of the cultural procedures utilized and by the point along the process chain at which sampling is being conducted. Cross-contamination of food products is a major factor that contributes to human illness. Campylobacter spp. can be sensitive to environmental conditions outside of an animal’s intestinal tract. While outbreaks of human campylobacteriosis have been associated with raw milk and untreated water, poultry meat, which is frequently contaminated with the organism, may be responsible for as much as 70% of sporadic campylobacteriosis. This chapter focuses on recent advantages in biological, chemical, and physical interventions to guard against the pathogen, and discriminative detection methods for confirmation and trace-back of contaminated products.

Clostridium perfringens is an anaerobic (microaerophilic), gram-positive, nonmotile, spore-forming, rod-shaped bacterium. C. perfringens is ubiquitous in the environment and is found in soil, dust, raw ingredients, such as spices used in food processing, and in the intestines of humans and animals. C. perfringens outbreaks usually result from improper handling and preparation of foods, such as inadequate cooling at the home, retail, or food service level, rarely involving commercial meat processors. This chapter discusses intrinsic and extrinsic factors that affect survival and growth in food products and contribute to outbreaks, and focuses on food processing operations that influence the numbers, spread, or characteristics. The presence of inhibitory agents in the products can affect germination of C. perfringens spores and may also affect the minimum growth temperatures for the germinated spores. Recent research has focused on combining traditional inactivation, survival, and growth-limiting factors at subinhibitory levels with emerging novel nonthermal intervention food preservation techniques using ionizing radiation, high hydrostatic pressure, or exposure to ozone. The ability of C. perfringens to cause food-borne illness and occasional associated outbreaks necessitates effective discriminatory detection methods for this pathogen in order to ensure reliable and confirmatory epidemiological screening of suspected foods. Many predictive growth models have been developed to accurately estimate C. perfringens survival following various types of food processing scenarios. The best strategy to control C. perfringens appears to be a hurdle approach combined with careful handling of foods to avoid temperature abuse.

In 1924 in Cambridge, England, what is now known as Listeria monocytogenes was first documented by E. G. D. Murray and colleagues (1926) as the causative agent of a septic illness, peripheral monocytosis, in laboratory rabbits. Between 1976 and 2002, there were 27 outbreaks of food-borne listeriosis reported worldwide, with about 2,900 cases and about 260 deaths (mortality rate of ca. 9.0%). L. monocytogenes is widespread in nature, being associated with plants, soil, water, sewage, feed, and animals raised as food. L. monocytogenes growth/survival at low temperatures requires maintenance of membrane fluidity for appropriate enzymatic activity and transport of solutes across the membrane, as well as for structure stabilization of macromolecules, such as ribosomes, and/ or the uptake or synthesis of compatible cryoprotectant solutes, such as glycine betaine and carnitine. Control of L. monocytogenes is one of the most difficult challenges faced by manufacturers and handlers of ready-to-eat (RTE) meat, poultry, seafood, and dairy products. Traditional milk pasteurization times/temperatures are also considered generally adequate to eliminate the typically low levels of L. monocytogenes that may be present in raw milk. The only bacteriocin that has been granted the FDA approval is nisin, which is allowed for use in low-moisture/lowsalt pasteurized processed cheese. Many organic acids have generally recognized as safe (GRAS) status as food ingredients, and studies have shown their effectiveness under various conditions and in different foodstuffs.

This chapter focuses predominantly on staphylococcal food poisoning (SFP), another toxin-mediated illness. SFP, also known as staphylococcal gastroenteritis, is not an infection but is a foodborne intoxication caused by one or more staphylococcal enterotoxins (SEs). It appears to have been initially reported in 1894 as a family outbreak due to undercooked meat from a sick cow that became contaminated with Staphylococcus aureus. Staphylococci are often divided into coagulasepositive staphylococci and coagulase-negative staphylococci (CNS), on the basis of coagulase production or the presence of a coagulase gene. Staphylococcal osmotolerance is problematic for food safety since blocked growth of competing bacteria can result in a more likely overgrowth of S. aureus. Staphylococci are ubiquitous in air, dust, sewage, water, milk, and many foods and on food equipment, environmental surfaces, humans, and animals. SE production is affected by temperature, pH, and osmotic conditions and is also regulated at the molecular level. The oligonucleotideoligosaccharide- binding fold containing a β-barrel structure capped with an α-helix comprises much of one domain and is present in other toxins including cholera toxin, pertussis toxin, and Shiga toxin. Recent studies demonstrated that staphylococcal pathogenicity island (SaPI) is mobilized by staphylococcal bacteriophages and endogenous prophages, suggesting that SE genes might be transferred horizontally. The identification of novel toxins and improved methods of detection have also indicated that SE production is much more widespread than originally thought and includes species of staphylococci other than S. aureus.

The primary pathogens in the genus Vibrio are Vibrio parahaemolyticus, V. cholerae, and V. vulnifius. Other Vibrio species that can cause human disease include V. hollisae, V. alginolyticus, V. damsela, V. mimicus, V. fluvialis , V. metschnikovii, V. furnissii, V. cincinnatiensis, and V. carchariae, but reported cases are either relatively rare or do not involve food-borne transmission. Nonepidemic V. cholerae is also responsible for occasional seafood-borne disease in the United States and can be distinguished from epidemic disease by the severity of symptoms, the serogroups of associated strains, and the capacity for global spread. V. vulnificus is the most common cause of serious wound infections associated with Vibrio species, and these infections may result from exposure of breached skin surface to seawater or contaminated seafood handling. This chapter talks about intrinsic factors and extrinsic factors. Comparison of heat resistances for pathogenic vibrios showed that D55 values varied among species. V. parahaemolyticus (D55 of 1.75 min) was considerably more resistant compared to V. vulnificus and V. cholera. Approved and validated treatments include high hydrostatic pressure, pasteurization (heat shock), and individual quick freezing. These treatments are generally used in combination with approved transport and storage practices involving icing, refrigeration, and/or freezing. Greater understanding of the role of the bacteria in estuarine ecosystems and the risks associated with environmental, bacterial, and host factors is crucial for control and safety of seafood products.

This chapter covers the bacterial pathogens that include: Aeromonas, Arcobacter, Helicobacter, Mycobacterium, Plesiomonas, and Streptococcus and attempts to provide a concise, thorough overview of the significance, characteristics, and food safety concerns involving each pathogen. Aeromonas species are indicated as important human pathogens causing gastrointestinal and other infections in healthy and immunocompromised hosts. “Aerolysins” are toxins produced by some Aeromonas species that have hemolytic, enterotoxic, and cytolytic activity. The importance of aeromonads as pathogens of food-borne origin dates back to the 1950s, following their isolation from humans. Campylobacter spp. have been identified as the leading cause of bacterial food-borne diarrheal illness in humans. The genus Helicobacter was created in 1989 and includes about 23 recognized species. Transmission of Helicobacter pylori through foods leading to incidences of food-borne illness has been speculated but not demonstrated to date. In both humans and animals, Mycobacterium avium subsp. paratuberculosis is thought to exist in protoplast form, which makes identification by acid-fast staining (Ziehl-Neelsen method) untenable. Plesiomonas shigelloides produces a heat-stable enterotoxin, but based on a lack of consistent in vitro and in vivo evidence, the species is thought to possess low pathogenicity. Earlier in the 20th century, there were efforts to reduce the incidence of food-borne illness due to Streptococcus species. While there are still occasional cases of illness reported, the intervention methods put in place have greatly reduced the likelihood of food-borne illness due to members of this genus.

This chapter focuses primarily on those viruses for which food-borne transmission is well documented (hepatitis A virus (HAV) and noroviruses (NoV)), and therefore, control in food production, processing, and preparation is considered relevant. General information about structural, molecular, and environmental properties of those viruses specifically implicated in food-borne illness is detailed. Three types of food commodities are usually associated with viral disease outbreaks, those being (i) molluscan shellfish contaminated during production; (ii) fresh produce items contaminated during production, harvesting, or packing; and (iii) prepared foods contaminated during preparation. Poor personal hygiene practices of infected food handlers provide the source of contamination for prepared foods. This appears to be the most important factor influencing the food-borne transmission of enteric viruses, as U.S. Centers for Disease Control and Prevention (CDC) data indicate that 50 to 95% of viral food-borne disease outbreaks are attributable to poor personal hygiene of infected food handlers. In Europe, researchers compiled data from 10 surveillance systems in the European Foodborne Virus Network, finding NoV to be the cause of >85% of all acute nonbacterial outbreaks of gastroenteritis reported between 1995 and 2000. Intrinsic and extrinsic parameters commonly used by food processors include manipulation of temperature (cooking and heating, freezing, and refrigeration), water activity, pH, gaseous environment, natural and intentionally added inhibitors, and the presence of competitive microflora. Our understanding of the importance of enteric viruses in the overall burden of food-borne disease has increased dramatically over the past 20 years.

This chapter discusses the formation and degradation of biogenic amines, their occurrence in foods, their significance in food safety, their potential use as quality indicators, and the available methods for their determination. The main mechanism of biogenic formation is the decarboxylation of free amino acids by specific enzymes of microbial origin, which leads to the production of amines. The concentration of available free amino acids plays a fundamental role in the formation of amines in foods, in that they are their precursors and, moreover, constitute a substrate for microbial growth. Histamine-producing microorganisms include a large number of strains from various species and families, isolated from a variety of food products. Pseudomonads are the most commonly reported bacterial group for putrescine and cadaverine formation. The addition of preservatives to foods influences the microbial population dynamics and, consequently, the production of biogenic amines. The physiological effects of tyramine include peripheral vasoconstriction, increased cardiac output, increased respiration, elevated blood glucose, and release of norepinephrine. In addition to being slightly toxic itself, tyramine reacts with monoamine oxidase (MAO) inhibitor (MAOI) drugs, giving rise to hypertensive crisis. MAO is an enzyme in the digestive tract that catalyzes the oxidative deamination of a variety of neurotransmitters as well as the monoamines of dietary origin, i.e., dopamine, phenyletylamine, serotonin, and tyramine. MAO has two subtypes, MAO-A and MAO-B, which can be found on the short arm of the X chromosome and seem to be derived from a duplication of a common ancestral gene.

This chapter focuses on a specific wheat allergen and the appropriate methods for its detection in food products to demonstrate the complexity of analytical challenges that the scientific community is facing. Such challenges apply to food allergen detection in general. In this chapter, the Osborne nomenclature is used because it is most cited among immunoassay developers. In the U.S. market, gluten commercial kit providers include Neogen Corp., whose enzyme-linked immunosorbent assay (ELISA) system based on two monoclonal antibody (mAbs), for rye and wheat, and Morinaga, whose ELISA uses polyclonal antibodies for wheat proteins. The gluten test is used to demonstrate the complexity of detection methods for food allergens in processed samples and the uncertainties and implications of results. Gluten content in foods can be evaluated by either commercial ELISA kits or lateral flow devices (LFD). Four gluten levels were evaluated in this study (50, 25, 10, and 5 ppm gluten) in two different matrices, gluten-free high-fiber bread mix and matrix-free 60% ethanol. The R-Biopharm kit results can be used to illustrate the problems associated with the selection and use of reference materials. The authors have used NIST gluten SRM 8418 to evaluate the kits, and have provided a detailed discussion on the characteristics of R5 (R-Biopharm) and Skerritt (Tepnel) monoclonal Abs (mAbs) based on findings and published information. The detection of gluten proteins is quite difficult due to the complexity of the targets and specificities of the Ab.

In this chapter, the use of the term residue relates primarily to the trace amounts of materials found in foods of various origins resulting from the application of chemicals during production. In order to focus on the type of sampling and the underlying statistical basis, the approach used for estimating the incidence of residues in animal products is presented in this chapter. The primary focus of the enforcement analytical work was related to antibiotic/antimicrobial assays and accounted for >99% of the samples. In sheep the incidence is 2.2%, approximately 2.5 times that reported in 2004, while in goats the 2005 frequency is 43% of that of 2004. A common acrylamide-forming reaction occurs when naturally occurring sugars react with asparagine at elevated temperatures. The most important point to be made is that acrylamide is considered a potential carcinogen and that the levels found in foods can be reduced during food preparation. The administration of the pesticide regulatory system is quite complex. Pesticide residues were not detected in 62.7% of domestic and 71.8% of imported samples. Of those samples, 2.4% of domestic and 6.1% of imported samples had violative levels. The problems with the U.S. food supplies are not the overall-occurring chemical residues but more likely the wide variety of bacteriological residues occurring from processing and distribution systems.

Prions are generally regarded as the most reliable markers of so-called prion diseases. With the advent of the protein-only concept of the transmission of prion diseases, which is predicated on the idea that prions are the infectious agents causing prion disease, and with the 1997 Nobel Prize for Medicine awarded for the prion protein concept, prions have firmly asserted their inclusion in the club of infectious agents, along with bacteria, viruses, fungi, and parasites, the club's traditional members. The essence of the protein-only theory is that the abnormal prion enters the organism and converts the host’s normal prions into abnormal prions. The efficiency of the conversion is influenced by the degree of amino acid sequence homology between the two PrPs involved, so that the greater the degree of homology, the greater the rate of conversion. Several studies suggest that white blood cells may be involved in the process. It was observed that gut-associated macrophages initially remove transmissible spongiform encephalopathy (TSE) infectivity, but once their ability to remove the infectivity completely is overcome, macrophages may facilitate the spread of infectivity. Regulatory measures introduced to prevent the spread of TSEs to humans and other animal species are likely to stay. It is possible, however, that more research and better tests will more adequately characterize risk, which in turn may lead to a gradual loosening of some of the currently existing restrictions following a carefully performed risk assessment.

This chapter discusses the interventions applied during harvest of different food commodities. Current interventions applied in order to reduce hazards during the harvesting of beef cattle include physical and chemical treatments. Chemical dehairing is a decontamination process that involves the removal of hair from animals by using a sodium sulfide solution followed by a rinse with a hydrogen peroxide solution and water prior to dehiding. The approved methods to meet this requirement are knife trimming and vacuuming beef carcasses with hot water or steam, when such contamination is less than 1 square inch. Steam pasteurization treatments have been used in meat processing. Several chemicals with antimicrobial activity have been tested for usefulness as carcass sanitizers. The steps in poultry slaughter for which contamination is most likely to occur are the scalding and defeathering points. High-pressure processing is an alternative treatment used to eliminate pathogens from foods with no thermal pasteurization and extend the shelf life of foods. Some studies developed to test the effectiveness of radiation in decontamination of seafood have demonstrated that doses of 3 kGy or below are enough to eliminate pathogenic bacteria. There are multiple interventions for hazard control during harvest that are capable of reducing pathogenic bacteria in foods of different origin. Overall, the proper use of approved interventions/ decontamination methods will continue to enhance the safety of the food supply and allow producers to meet the demands and expectations of the consumers.

This chapter deals with biological hazards and, more specifically, viral and bacterial pathogens. It provides an overview of the hazards and risks associated with the presence and transmission of viral and bacterial pathogens in retail-handled ready-to-eat (RTE) foods, and discusses potential control interventions. The limited number of reports of outbreaks of viral infections originating from retail-handled foods should not be considered as evidence that viral pathogens are not of importance at retail. Assurance for hazard control on incoming products can be derived from purchase specifications and supplier audits. Furthermore, good hygiene practices (GHP) including regular and thorough hand washing and use of barriers to bare-hand contact with RTE foods, such as gloves, deli wraps, and utensils, can contribute considerably to the interruption of both of the food handler-associated routes of pathogen transmission. Indeed, GHP appear to be the most important hazard control intervention, with regard to food handlers that may be asymptomatic carriers of pathogenic organisms. Effective risk communication, education, and training of food workers, supervisors, and managers are considered prerequisites for the successful implementation of hazard control measures at retail.

The continued presence of pathogenic microorganisms and their toxins in food and drinking water has necessitated the ongoing need for newer, more sensitive and robust analytical systems capable of rapid detection of these contaminants in complex samples. The ideal detection method should be capable of rapidly detecting and confirming the presence of food-borne pathogenic microorganisms directly from complex samples with no false-positive or false-negative results. Rapid detection methods including immunological detection, cell/tissue-based assays and nucleic acid-based assays have been discussed in this chapter. Conventional culture techniques continue to be the gold standard for the isolation, detection, and identification of target pathogens. These methods increase detection times by hours to days, causing preliminary test results to be delayed. These assays are defined as affinity, cell/ tissue, and nucleic acid technologies. Antibody-based detection systems are still considered to be the gold standard of affinity-based testing methods. Aptamers offer several advantages over the use of antibodies in the identification of food-borne microorganisms and toxins. Any microorganism that contains DNA or RNA can be detected using nucleic acid-based assays, but a limitation of these diagnostics is their inability to detect protein-based agents of disease, such as toxins or prions.

Food safety management can be viewed from different levels: broadly from the perspective of government food safety management systems or more narrowly from that of industry-wide food safety management systems or that of food safety management within an individual establishment. The role of the food industry is to provide safe foods; industry is responsible for establishing food safety management systems that ensure that foods present a minimal risk to the consumer. Risk analysis allows the development of risk-based metrics, such as food safety objectives (FSOs), performance objectives (POs), and performance criteria (PCs), which are addressed in this chapter. Seven hazard analysis and critical control point (HACCP) principles form the framework of a systematic approach to ensure the determination and control of significant food safety hazards associated with a product and process. Monitoring is the first line of defense in the preventive HACCP system. In addition to those related to GMP or sanitation regulations, prerequisite programs can include other programs, such as ingredient specifications, consumer complaint management, glass control programs, allergen management programs, microbiological monitoring of the plant environment, ingredient-to-product traceability programs, and supplier approval programs. PCs can easily be translated into product or process criteria by industry or by government. In integrating a farm-to-fork food safety system, there will be a variety of metrics at points along the food chain.