The Microbiology of Safe Food

Preface to third edition 12

Preface to second edition 14

Preface to first edition 16

Chapter 1 : Foodborne infections 18

1.1 The microbial world and its relationship to food 19

1.2 Origins of safe food production 25

1.3 Overview of foodborne illness 27

1.4 Public perception of safe food 32

1.5 Causes of foodborne illness 36

1.6 Food poisoning due to common food commodities 39

1.6.1 Milk and milk products 39

1.6.2 Meat products 40

1.6.3 Fresh produce 41

1.6.4 Low-water activity (a_w) and low-moisture foods 41

1.7 Host-related issues 43

1.8 Hygiene hypothesis 46

1.9 Chronic sequelae following foodborne illness 47

1.10 The size of the foodborne illness problem 49

1.11 The cost of foodborne diseases 60

1.12 Changes in antimicrobial resistance of foodborne pathogens 65

1.12.1 Bacterial antibiotic resistance in agriculture and aquaculture 65

1.12.2 Antibiotics of concern and resistance mechanisms 69

1.12.3 Polymixin and plasmid-encoded colistin resistance 71

1.12.4 Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) 73

1.13 Food safety following natural disasters, and conflict 74

1.14 Food microbiology foodborne diseases and climate change 75

Chapter 2 : Basic aspects 77

2.1 The human intestinal tract 77

2.2 The normal human intestinal flora 78

2.3 Host resistance to foodborne infections 83

2.4 Bacterial cell structure 86

2.4.1 Morphology 86

2.4.2 Cell membrane structure and the Gram stain 86

2.4.3 Lipopolysacharide (LPS, O antigen) 88

2.4.4 Flagella (H antigen) 89

2.4.5 Capsule (K- and Vi-antigen) 90

2.5 Bacterial toxins and other virulence determinants 91

2.5.1 Bacterial endotoxins and exotoxins 91

2.5.2 Pathogenicity islands 97

2.5.3 Bacterial toxins encoded in bacteriophages 99

2.6 Microbial growth cycle 99

2.7 Death kinetics 101

2.7.1 Expressions 101

2.7.2 Decimal reduction times (D-values) and z values 102

2.8 Factors affecting microbial growth 105

2.8.1 Intrinsic and extrinsic factors affecting microbial growth 106

2.8.2 Water activity 106

2.8.3 pH 108

2.8.4 Temperature 109

2.8.5 Interplay of factors affecting microbial growth in foods 109

2.9 Microbial response to stress 110

2.9.1 General stress response (GSR) 111

2.9.2 pH stress 113

2.9.3 Heat-shock 114

2.9.4 Cold-shock 116

2.9.5 Osmotic shock 117

2.10 Predictive modeling 118

2.10.1 Predicting modeling development 119

2.10.2 Primary models and the Gompertz and Baranyi equations 121

2.10.3 Secondary models 124

2.10.4 Tertiary models 125

2.10.5 Application of predictive microbial modeling 126

Chapter 3 : Food preservation and spoilage organisms 128

3.1 Spoilage microorganisms 128

3.1.1 Spoilage microorganisms 130

3.1.2 Spoilage of diary products 133

3.1.3 Spoilage of meat products 134

3.1.4 Fish spoilage 136

3.1.5 Egg spoilage 136

3.1.6 Cereals and grain 137

3.2 Shelf-life indicators 138

3.3 Methods of preservation and shelf-life extension 141

3.4 Preservatives 143

3.4.1 Organic acids 144

3.4.2 Hydrogen peroxide and lactoperoxidase system 144

3.4.3 Chelators 146

3.4.4 Non-acidic preservatives 146

3.4.5 Preservation due to weak acids and low pH 148

3.4.6 Biopreservatives 148

3.5 Physical methods of preservation 150

3.5.1 Preservation by heat-treatment 150

3.5.2 High pressure treatment. 152

3.5.3 Ohmic heating and radio frequency 154

3.5.4 Pulsed electric fields 154

3.5.5 Ultrasound 155

3.5.6 Intense light pulse 155

3.5.7 Food irradiation 156

3.5.8 Cold plasma and low-energy electrons for food surface decontamination 163

3.6 Packaging 164

3.6.1 Reduced oxygen packaging, modified atmosphere packaging and active packaging 164

3.6.2 Antimicrobial packaging and nanotechnology 167

3.7 Fermented food products 170

3.7.1 Fermented milk products 170

3.7.2 Fermented meat products 174

3.7.3 Fermented vegetables 175

3.7.4 Fermented protein foods; shoyu and miso 175

3.8 Organisms involved in the production of fermented foods 175

3.8.1 Lactic acid bacteria 177

3.8.2 Bifidobacterium species 184

3.8.3 Other organisms 187

3.9 Functional foods: probiotics and gut modulation 188

3.9.1 Qualified Presumption of Safety (QPS) and Generally Regarded as Safe (GRAS) 188

3.9.2 Functional foods and probiotics 190

3.9.3 Probiotic studies 194

3.9.4 Novel organisms – modulation of gut microbiota 196

Chapter 4 : Bacterial foodborne pathogens 199

4.1 Indicator organisms 200

4.1.1 Coliforms 202

4.1.2 Enterobacteriaceae 203

4.1.3 Enterococci 203

4.1.4 Bacteriophages 204

4.2 Campylobacter jejuni, C. coli and C. lari 205

4.2.1 General description 205

4.2.2 Campylobacter infections 207

4.2.3 Campylobacter jejuni typing 209

4.2.4 Virulence factors 212

4.2.5 Whole genome sequence analysis 216

4.2.6 Sources and control of Campylobacter jejuni 218

4.3 Salmonella serovars 221

4.3.1 General description 221

4.3.2 Salmonella serotypes 223

4.3.3 Infections caused by Salmonella serovars 227

4.3.4 Virulence factors of Salmonella serovars 232

4.3.5 Whole genome analysis 233

4.3.6 Sources and control of Salmonella serovars 235

4.3.7 Salmonella serovar outbreaks 238

4.4 Pathogenic Escherichia coli 245

4.4.1 General description 245

4.4.2 E. coli pathovars 245

4.4.3 Infections caused by E. coli pathovars 247

4.4.4 Virulence factors 254

4.4.5 Whole genome analysis 255

4.4.6 Sources and control of E. coli pathovars 257

4.4.7 Outbreaks caused by E. coli pathovars 259

4.5 Shigella dysenteriae and Sh. sonnei 267

4.5.1 General description 267

4.5.2 Shigellosis 268

4.5.3 Virulence factors 268

4.5.4 Shigella sonnei outbreak 270

4.6 Cronobacter species 271

4.6.1 General description 271

4.6.2 Infections due to Cronobacter species 272

4.6.3 Identification and typing methods for Cronobacter species 273

4.6.4 Virulence factors 274

4.6.5 Sources and control of Cronobacter species 276

4.6.6 Cronobacter outbreaks 279

4.7 Vibrio cholerae, V. parahaemolyticus , and V. vulnificus 280

4.7.1 General description 280

4.7.2 Infections due to Vibrio species 281

4.7.3 Virulence factors 284

4.7.4 Sources and control 286

4.8 Brucella melitensis, Br. abortus and Br. suis 287

4.8.1 General description 287

4.8.2 Brucellosis 288

4.9 Yersinia enterocolitica 289

4.9.1 General description 289

4.9.2 Yersiniosis 289

4.9.3 Sources and control 291

4.9.4 Outbreaks due to Y. enterocolitica 292

4.10 Aeromonas hydrophila, A. caviae and A. sobria 293

4.10.1 General description 293

4.10.2 A. hydrophila gastroenteritis 293

4.10.3 Sources and control 295

4.11 Pleisiomonas shigelloides 296

4.11.1 General description 296

4.11.2 Plesiomonas infections 296

4.11.3 Sources and control 297

4.12 Listeria monocytogenes 298

4.12.1 General description 298

4.12.2 Listeriosis 299

4.12.3 Lineages and typing L. monocytogenes 301

4.12.4 Virulence factors 304

4.12.5 Whole genome sequencing of L. monocytogenes 309

4.12.6 Sources and control of L. monocytogenes 312

4.12.7 L. monocytogenes outbreaks 315

4.13 Staphylococcus aureus 324

4.13.1 General description 324

4.13.2 Infections associated with St. aureus 325

4.13.3 Virulence factors 326

4.13.4 Sources and control 326

4.14 Clostridium perfringens 328

4.14.1 General description 328

4.14.2 Cl. perfringens infections 328

4.14.3 Sources and control 330

4.15 Clostridium botulinum 330

4.15.1 General description 330

4.15.2 Cl. botulinum intoxication 331

4.15.3 Sources and control 332

4.16 Bacillus cereus group 332

4.16.1 General description 332

4.16.2 B. cereus foodborne infections 334

4.16.3 Virulence traits 335

4.16.4 Sources and control 336

4.17 Enterococcus and Streptococcus species. 338

4.17.1 General description 338

4.17.2 Enterococcus faecalis and E. faecium 339

4.17.3 Streptococcus pyogenes, Group A streptococci 340

4.17.4 Virulence traits 341

4.18 Emerging and uncommon foodborne pathogens 342

4.18.1 Arcobacter genus 346

4.18.2 Campylobacter concisus 349

4.18.3 Enteroaggregative Escherichia coli (EAEC), E. coli O55 and E. coli O26 350

4.18.4 Escherichia albertii 355

4.18.5 Providencia alcalifaciens 358

4.18.6 Clostridium difficile 359

4.18.7 Mycobacterium paratuberculosis 363

4.18.8 Acinetobacter species 364

4.18.9 Nanobacteria 365

Chapter 5: Foodborne pathogens: viruses, toxins, parasites and prions 366

5.1 Foodborne viruses 366

5.1.1 Norovirus 370

5.1.2 Hepatitis A 376

5.1.3 Hepatitis E. 381

5.1.4 Rotaviruses 382

5.1.5 Small round viruses, astroviruses, Sapporo-like viruses, adenoviruses and parvoviruses 384

5.1.6 Human enteroviruses 386

5.2 Seafood and shellfish poisoning 387

5.2.1 Ciguatera poisoning 388

5.2.2 Scombroid poisoning 388

5.2.3 Paralytic shellfish poisoning 389

5.2.4 Diarrhaeic shellfish poisoning 389

5.2.5 Neurotoxic shellfish poisoning 390

5.2.6 Amnesic shellfish poisoning 390

5.3 Foodborne parasites: eucaryotes 391

5.3.1 Toxoplasma gondii 392

5.3.2 Taenia saginata and T. solium 393

5.3.3 Echinococcus multilocularis and E. granulosus 393

5.3.4 Cyclospora cayetanensis 394

5.3.5 Cryptosporidum parvum 396

5.3.6 Anisakis simplex 398

5.3.7 Trichinella spiralis 398

5.4 Mycotoxins 399

5.4.1 Aflatoxins 400

5.4.2 Ochratoxins 402

5.4.3 Fumonisins 403

5.4.4 Zearalenone 403

5.4.5 Trichothesenes 403

5.4.6 Prions and transmissible spongiform encephalopathies 404

Chapter 6 : Methods of detection and characterisation 407

6.1 Prologue 407

6.2 Conventional methods 414

6.2.1 Culture media 416

6.2.2 Sublethally injured cells 419

6.2.3 Viable but nonculturable bacteria (VBNC) 421

6.3 Rapid sampling methods 422

6.3.1 Sample preparation 423

6.3.2 Separation and concentration of target 423

6.4 Rapid end detection methods 426

6.4.1 ATP bioluminescence techniques and hygiene monitoring. 427

6.4.2 Protein detection 429

6.4.3 Flow cytometry 429

6.4.4 Biosensors 430

6.4.5 Impedance (conductance) microbiology 433

6.5 DNA-based molecular typing and proteomic methods 435

6.5.1 Polymerase chain reaction (PCR) 436

6.5.2 Microarrays 439

6.5.3 Loop-mediated isothermal amplification (LAMP) technique 441

6.5.4 Pulsed-field gel electrophoresis (PFGE) 442

6.5.5 Restriction fragment length polymorphism (RFLP) 444

6.5.6 Amplified fragment length polymorphism (AFLP) 445

6.5.7 Random amplification of polymorphic DNA (RAPD) 445

6.5.8 Repetitive-element polymerase chain reaction (Rep-PCR) 446

6.5.9 Nucleic acid sequence-based amplification (NASBA) 447

6.5.10 Multiple locus variable number tandem repeat analysis (MLVA) 447

6.5.11 PCR-probe based serotyping 449

6.5.12 Ribotyping 450

6.5.13 MALDI-TOF 450

6.6 Identification and typing methods based on high-throughput DNA sequencing 451

6.6.1 Conventional 7-loci multilocus sequence typing (MLST) 452

6.6.2 Genome sequence based MLST 454

6.6.3 (Plate 19)-cas array typing 457

6.6.4 Single nucleotide polymorphism-based (SNP) analysis 458

6.7 Specific detection procedures and acceditation 459

6.7.1 Aerobic plate count 460

6.7.2 Salmonella serovars 461

6.7.3 Campylobacter 465

6.7.4 Enterobacteriaceae and E. coli 467

6.7.5 Pathogenic E. coli, including E. coli O157:H7 470

6.7.6 Shigella species 472

6.7.7 Cronobacter genus 473

6.7.8 Aeromoas species 474

6.7.9 Arcobacter species 475

6.7.10 Listeria monocytogenes 476

6.7.11 Staphylococcus aureus 479

6.7.12 Clostridium perfringens 481

6.7.13 Bacillus cereus, B. subtilis and B. licheniformis 482

6.7.14 Mycotoxins 483

6.7.15 Viruses 483

Chapter 7 : Microbiological Criteria 485

7.1 Background to microbiological criteria and end-product testing 485

7.2 International Commission on Microbiological Specifications for Foods (ICMSF) 486

7.3 Codex Alimentarius principles for the establishment and application of microbiological criteria 488

7.4 Sampling Plans 491

7.5 Variables plans 494

7.6 Attriibutes sampling plan 497

7.6.1 Two class plan 497

7.6.2 Three class plan 498

7.7 Principles 499

7.7.1 Defining a ‘lot’ of food 499

7.7.2 Sample unit number 499

7.7.3 Operating Characterisitic curve 500

7.7.4 Producer risk and consumer risk 502

7.7.5 Stringency of two and three class plans, setting n and c. 502

7.7.6 Setting the values for m and M 504

7.8 Microbiological Limits 505

7.8.1 Definitions 505

7.8.2 Limitations of microbiological testing 506

7.8.3 Examples of sampling plans 506

7.9 Implemented microbiological criteria 508

7.9.1 Microbiological criteria in the European Union 508

7.9.2 EU Directives specifying microbiological standards for foods 509

7.10 UK guidelines for ready-to-eat foods 510

Chapter 8 : Hygienic Production Practices 512

8.1 Contribution of food handlers to foodborne illness 512

8.2 Personnel hygiene and training 513

8.3 Cleaning 517

8.4 Detergents and disinfectants 519

8.5 Microbial biofilms 520

8.5.1 Microbial biofilm formation 520

8.5.2 Bacterial biofilm induction 523

8.5.3 Biofilm removal and control 524

8.6 Assessment of cleaning and disinfection efficiency 527

Chapter 9 : Food Safety Management Tools 530

9.1 The manufacture of hygienic food 530

9.2 Microbiological safety of food in world trade 537

9.3 Consumer pressure affect on food processing 539

9.4 The management of hazards in food which is in international trade 540

9.5 Hazard Analysis Critical Control Point (HACCP) 542

9.6 Prerequisite Programme 544

9.7 Outline of HACCP 545

9.7.1 Food hazards 546

9.7.2 Preparation for HACCP 547

9.7.3 Principle 1: Hazard analysis 548

9.7.4 Principle 2: Critical Control Points 549

9.7.5 Principle 3: Critical Limits 550

9.7.6 Principle 4: CCP monitoring 550

9.7.7 Principle 5: Corrective actions 551

9.7.8 Principle 6: Verification 551

9.7.9 Principle 7: Record Keeping 553

9.8 Microbiological Criteria and HACCP 554

9.9 Microbiological hazards and their control 554

9.9.1 Sources of microbiological hazards 554

9.9.2 Temperature control of microbiological hazards 556

9.9.3 Non-temperature control of microbiological hazards 559

9.10 HACCP plans 559

9.10.1 Production of pasteurised milk 560

9.10.2 Swine slaughter in the abattoir 560

9.10.3 Chilled food manufacture 561

9.10.4 Generic models 561

9.11 Good Manufacturing Practice (GMP) and Good Hygiene Practice (GHP) 562

9.12 Quality Systems 563

9.13 Total Quality Management (TQM) 564

Chapter 10 : Microbiological Risk Assessment 566

10.1 Risk analysis and microbiological risk assessment 566

10.2 Origin of microbiological risk assessment 569

10.3 Microbiological risk assessment – an overview 572

10.4 Microbiological Risk Assessment – structure 577

10.4.1 Risk Assessment 579

10.4.2 Risk Management 581

10.4.3 Risk Communication 581

10.5 Risk Assessment 581

10.5.1 Statement of purpose 582

10.5.2 Hazard Identification 583

10.5.3 Exposure Assessment 584

10.5.4 Hazard Characterization 588

10.5.5 Dose-response assessment 591

10.5.6 Dose-response models 594

10.5.7 Dose and infection 600

10.5.8 Risk Characterization 606

10.5.9 Production of a formal report 608

10.5.10 Triangular distributions and Monte Carlo simulation 608

10.6 Risk Management 610

10.6.1 Risk assessment policy 614

10.6.2 Risk Profiling. 614

10.7 Food Safety Objectives 615

10.8 Risk Communication 618

10.9 Future developments in microbiological risk assessment 621

10.9.1 International methodology and guidelines 622

10.9.2 Risk assessment database 622

10.9.3 Training courses and use of resources 624

Chapter 11 : Application of Microbiological Risk Assessment 626

11.1 Salmonella serovars 627

11.1.1 S. Enteritidis in shell eggs and egg products 627

11.1.2 Hazard identification and hazard characterization of Salmonella in broilers and eggs 630

11.1.3 Exposure assessment of Salmonella serovars in broilers 632

11.1.4 Salmonella serovars in cooked chicken 635

11.1.5 Salmonella serovars in cooked patty 637

11.1.6 Poultry FARM 638

11.1.7 Domestic and sporadic human salmonellosis 638

11.2 Campylobacter 639

11.2.1 C. jejuni risk from fresh chicken 639

11.2.2 Risk profile for pathogenic species of Campylobacter in Denmark 642

11.2.3 Risk assessment of C. jejuni in broilers 643

11.2.4 Campylobacter fluoroquinolone resistance 644

11.3 L. monocytogenes 648

11.3.1 L. monocytogenes hazard identification and hazard characterization in ready-to-eat foods 648

11.3.2 L. monocytogenes exposure assessment in ready-to-eat foods 649

11.3.3 Relative risk of L. monocytogenes in selected ready-to-eat foods 651

11.3.4 L. monocytogenes in EU trade 652

11.3.5 L. monocytogenes in meat balls 654

11.3.6 Listeriosis from ready-to-eat meat products 655

11.4 E. coli O157 655

11.4.1 E. coli O157:H7 in ground beef 655

11.5 B. cereus 658

11.5.1 B. cereus risk assessment 658

11.6 V. parahaemolyticus 659

11.6.1 Public health impact of V. parahaemolyticus in raw molluscan shellfish 659

11.7 Cronobacter species and Salmonella in powdered infant formula 661

11.8 Viral risk assessments 663

11.8.1 Viral contamination of shellfish and coastal waters 664

Chapter 12 : International control of microbiological hazards in foods; regulations and authorities666

12.1 Control of foodborne pathogens 666

12.1.1 Control of Salmonella serovars in poultry 666

12.1.2 Control of E. coli pathovars and Salmonella serovars in fresh produce 670

12.1.3 Control of pathogens in low-moisture foods 673

12.2 World Health Organisation, global food security from accidental and deliberation contamination 677

12.3 Regulations in international trade of food 682

12.4 Codex Alimentarius Commission (CAC) 684

12.5 Sanitary and Phytosanitary measures (SPS), Technical Barriers to Trade (TBT) and the World Health Organisation (WHO) 685

12.6 European Union legislation 688

12.7 International food safety agencies 691

12.7.1 European Food Safety Authority (EFSA) 692

12.7.2 Food authorities in the United States 693

Chapter 13 : Surveillance and foodborne outbreak investigation 697

13.1 Surveillance programs 697

13.1.1 International Food Safety Authorities Network (INFOSAN) 700

13.1.2 Surveillance systems in the United States (FoodNet) 700

13.1.3 PulseNet International 703

13.1.4 European Centre for Disease Prevention and Control (ECDC) and european surveillance for salmonellosis and shiga toxin-producing E. coli (STEC) 706

13.1.5 European Food-borne Viruses in Europe network (FBVE) 707

13.1.6 Rapid Alert System for Food and Feed (RASFF) 708

13.1.7 Global Salm-Surv (GSS) 711

13.1.8 Surveillance of ready-to-eat foods in the United Kingdom 712

13.2 Outbreak investigations 713

13.2.1 Preliminary outbreak investigation 716

13.2.2 Case definition and data collection 717

13.2.3 Data collation and interpretation 719

13.4 Social media, crowd sourcing and reporting food poisoning cases 724

13.5 Mobile phones and food safety 725

13.6 Food terrorism and biocrimes 726

Chapter 14 : Whole genome sequencing, microbiomes and genomic epidemiology 734

14.1 High-throughput DNA sequencing 734

14.2 Microbiome analysis 739

14.3 Genomic epidemiology 742

14.4.1 Whole genome sequencing for microbial source tracking 742

14.4.2 Genome Trakr network (US) 746

14.4.3 NCBI Pathogen Detection 746

14.4.4 Center for Genomic Epidemiology (Denmark) 747

14.5 Key outbreaks investigated using genomic epidemiology 747

14.5.1 Ready-to-eat meat products L. monocytogenes outbreak, Canada 2008 747

14.5.2 E. coli O104:H4 outbreak, Germany 2011 749

14.5.3 C. jejuni outbreak investigations 753

14.5.4 Salmonella Enteritidis in eggs, European outbreak 2014 755

14.5.5 Multinational outbreak of Salmonella Agona through infant formula contamination, 2017 757

14.5.6 RetrospectiveCronobacter sakazakii neonatal intensive care unit outbreak, France 1994 760

14.5.7 L. monocytogenes ST6, polony sausages, South Africa, 2017-18 762

Glossary of terms : 765

List of abbreviations: 776

Food Safety Resources on the World Wide Web 777

References 789

Exploring food microbiology, its impact upon consumer safety, and the latest strategies for reducing its associated risks

As our methods of food production advance, so too does the need for a fuller understanding of food microbiology and the critical ways in which it influences food safety. The Microbiology of Safe Food satisfies this need, exploring the processes and effects of food microbiology with a detailed, practical approach. Examining both food pathogens and spoilage organisms, microbiologist Stephen J. Forsythe covers topics ranging from hygiene regulations and product testing to microbiological criteria and sampling plans.

This third edition has been thoroughly revised to cater to the food scientists and manufacturers of today, addressing such new areas as: 

• Advances in genomic analysis techniques for key organisms, including E. coli, Salmonella, and L. monocytogenes
• Emerging information on high-throughput sequencing and genomic epidemiology based on genomic analysis of isolates
• Recent work on investigations into foodborne infection outbreaks, demonstrating the public health costs of unsafe food production
• Updates to the national and international surveillance systems, including social media

Safe food for consumers is the ultimate goal of food microbiology. To that end, The Microbiology of Safe Food focuses on the real-world applications of the latest science, making it an essential companion for all those studying and working in food safety. 

Stephen J. Forsythe is former Professor of Microbiology at Nottingham Trent University, Nottingham, UK. He is currently visiting Professor to many universities and sits on a number of governmental advisory committees. He has many years of experience teaching food microbiology to university students and professionals within the food industry and government regulatory bodies.