Microbial Toxins in Dairy Products Society of Dairy Technology Series

Food-borne diseases, including those via dairy products, have been recognised as major threats to human health. The causes associated with dairy food-borne disease are the use of raw milk in the manufacture of dairy products, faulty processing conditions during the heat treatment of milk, post-processing contamination, failure in due diligence and an unhygienic water supply. Dairy food-borne diseases affecting human health are associated with certain strains of bacteria belonging to the genera of Clostridium, Bacillus, Escherichia, Staphylococcus and Listeria, which are capable of producing toxins, plus moulds that can produce mycotoxins such as aflatoxins, sterigmatocytin and ochratoxin.

Microbial Toxins in Dairy Products reviews the latest scientific knowledge and developments for detecting and studying the presence of these toxins in dairy products, updating the analytical techniques required to examine bacterial and mould toxins and the potential for contamination of milk as it passes along the food chain, i.e. from 'farm-to-fork'.

This comprehensive and accessible collection of techniques will help dairy processors, food scientists, technologists, researchers and students to further minimise the incidences of dairy food-borne illnesses in humans.

List of Contributors xi

Preface to the Technical Series xv

Preface xvi

1 Microbial Toxins – An Overview 1
R. Early and A.Y. Tamime

1.1 Introduction 1

1.2 Microbial toxins: modes of action 2

1.3 Bacterial toxins 4

1.3.1 Staphylococcal enterotoxins (SEs) 4

1.3.2 Bacillus cereus group enterotoxins 6

1.3.3 Clostridium botulinum nerotoxin 6

1.4 Mycotoxins 7

1.4.1 Background 7

1.4.2 General aspects of mycotoxins 7

1.4.3 Postscript on mycotoxins 13

1.5 Biogenic amines (BAs) 14

1.6 Conclusions 14

References 16

2 Incidences of Mould and Bacterial Toxins in Dairy Products 19
M.L.Y. Wan, N.P. Shah and H.I. El-Nezami

2.1 Background 19

2.2 Bacterial toxins 19

2.2.1 Emetic toxin produced by Bacillus cereus 20

2.2.2 Enterotoxins produced by Staphylococcus aureus 24

2.2.3 Botulinum neurotoxins produced by Clostridium botulinum 26

2.3 Mould toxins (mycotoxins) 33

2.3.1 Aflatoxins B1 and M1 34

2.3.2 Sterigmatocystin 42

2.3.3 Ochratoxin A 43

2.4 Other mycotoxins 47

2.5 Conclusion 49

References 50

3 Bacterial Toxins – Structure, Properties and Mode of Action 71
J.W. Austin

3.1 Background 71

3.2 Bacillus cereus toxins 72

3.2.1 Bacillus cereus emetic toxin 73

3.2.2 Bacillus cereus enterotoxins 74

3.2.3 Bacillus cereus haemolysin BL (Hbl) 76

3.2.4 Bacillus cereus non-haemolytic enterotoxin (Nhe) 76

3.2.5 Cytotoxin K (Cyt K) 77

3.3 Botulinum neurotoxin 77

3.3.1 Outbreaks of botulism caused by dairy products 78

3.3.2 Structure of botulinum neurotoxin 79

3.3.3 Mode of action of BoNTs 80

3.4 Staphylococcus aureus enterotoxin 80

3.5 Conclusions 83

References 83

4 Biogenic Amines in Dairy Products 94
V. Ladero, D.M. Linares, M. Pérez, B. del Rio, M. Fernández and M.A. Alvarez

4.1 Introduction 94

4.2 Biochemistry: biosynthesis pathways, enzymes and transporters 98

4.2.1 Tyramine 98

4.2.2 Histamine 99

4.2.3 Putrescine 100

4.2.4 Cadaverine, β-phenylethylamine, tryptamine 101

4.3 Biogenic amine-producing micro-organisms 101

4.3.1 Genes involved in the biosynthesis of biogenic amines 103

4.3.2 Is the production of BAs a strain- or species-dependent characteristic? 105

4.3.3 Physiological functions of BAs biosynthesis 106

4.4 Toxicological effects 107

4.4.1 Tyramine 108

4.4.2 Histamine 109

4.4.3 Putrescine and polyamines 110

4.4.4 Cadaverine, tryptamine and β-phenylethylamine 111

4.4.5 Recommended limits of BAs 111

4.5 Factors affecting BAs accumulation in dairy products 113

4.5.1 Presence of BAs-producing bacteria 113

4.5.2 Physiochemical factors 114

4.5.3 Technological factors 117

4.6 Other preventive methods 119

4.7 Conclusions 119

Acknowledgements 120

References 120

5 Contamination of Raw Milk: Sources and Routes Up to the Farm Gate 132
R. Early

5.1 Introduction 132

5.2 The concept of contamination 132

5.2.1 What does contamination mean to the concept of food? 134

5.2.2 Contamination and cow health 135

5.3 Sources of contamination 136

5.3.1 Biological contamination 136

5.3.2 Chemical contamination 142

5.3.3 Mycotoxins 147

5.3.4 Physical contamination 148

5.4 Conclusion 150

References 150

6 Milk Product Contamination After the Farm Gate 154
R. Early

6.1 Introduction 154

6.2 The significance of microbial contamination 154

6.2.1 Product spoilage 154

6.2.2 Food-borne illness 155

6.2.3 Microbial toxins 155

6.3 Factories, processes and people 157

6.4 Raw milk handling 158

6.5 Milk-processing and dairy products manufacture 160

6.5.1 Liquid milk and cream processing 161

6.5.2 Packing, storage, distribution and the retail environment 162

6.6 Buttermaking 164

6.7 Cheesemaking 165

6.8 Yoghurt 168

6.9 Milk powders 170

6.10 Evaporated milk and sweetened condensed milk 172

6.10.1 Evaporated milk 172

6.10.2 Sweetened condensed milk 175

6.11 Ice-cream 175

6.12 Hygiene, food safety management and cleaning-in-place (CIP) 177

6.13 Packaging, storage, distribution and the retail environment 178

6.14 Conclusions 180

References 180

7 Techniques for Detection, Quantification and Control of Bacterial Toxins 183
L. Ramchandran, A. Warnakulasuriya, O. Donkor and T. Vasiljevic

7.1 Introduction 183

7.2 Bacterial toxins 184

7.3 Control of toxins 186

7.4 Methods for identification and detection of microbial toxins 187

7.4.1 Traditional biological assays 189

7.4.2 Antibody and immunoassay 191

7.5 Conclusion 196

References 196

8 Techniques for Detection, Quantification and Control of Mycotoxins in Dairy Products 201
O. Donkor, L. Ramchandran and T. Vasiljevic

8.1 Introduction 201

8.2 Methods for detection and quantification of mycotoxins 203

8.2.1 Sample pre-treatment method 203

8.2.2 Liquid-liquid extraction 204

8.2.3 Supercritical fluid extraction 204

8.2.4 Solid phase extraction 204

8.3 Separation methods 206

8.3.1 Thin layer chromatography 206

8.3.2 High pressure liquid chromatography (HPLC) 208

8.3.3 Gas chromatography (GC) 210

8.3.4 Capillary electrophoresis (CE) 210

8.3.5 Biosensors 210

8.3.6 Enzyme-linked immmunosorbent assay (ELISA) method 212

8.3.7 Electrochemical immunoassay 214

8.3.8 Polymerase chain reaction (PCR)-based detection and quantification 215

8.4 Mathematical model (exposure assessment of mycotoxins in dairy milk) 216

8.5 Control of mycotoxin 217

8.5.1 Physical methods 218

8.5.2 Chemical methods 218

8.5.3 Biological methods 219

8.5.4 Activated carbon (AC) 219

8.6 Conclusion 220

References 220

9 Approaches to Assess the Risks/Modelling of Microbial Growth and Toxin Production 229
N. Murru, R. Mercogliano, M.-L. Cortesi, F. Leroy, R. Condoleo and M.F. Peruzy

9.1 Background on risk analysis 229

9.2 Focus on cheese risk assessment 231

9.2.1 Source of milk 231

9.2.2 Raw and/or heat-treated milk cheeses 231

9.2.3 Level of moisture in cheese 232

9.2.4 Methods of manufacture 232

9.2.5 Fat content 232

9.2.6 Maturation indices 232

9.2.7 Washed or mould cheeses 233

9.3 Staphylococcus aureus 233

9.3.1 Background 233

9.3.2 Staphylococcus aureus and production of staphylococcal eneterotoxins 234

9.3.3 Cheese production and hazard characterisation of Staphylococcus aureus 237

9.3.4 Cheesemaking conditions and exposure assessment of Staphylococcus aureus 238

9.3.5 Predictive modelling and risk assessment of Staphylococcus aureus and enterotoxin production in cheese 242

9.4 Escherichia coli 243

9.4.1 Hazard identification 243

9.4.2 Growth and inactivation 244

9.4.3 Hazard characterisation 244

9.4.4 Exposure assessment 246

9.4.5 Risk characterisation 248

9.5 Listeria monocytogenes 251

9.5.1 Hazard characterisation 251

9.5.2 Exposure assessment 253

9.5.3 Hazard characterisation 256

9.5.4 Risk characterisation 258

9.6 Cheese - chemical risk assessment 259

9.6.1 Background 259

9.6.2 Biogenic amines in cheese 260

9.6.3 Occurrence of biogenic amines in cheese: hazard and exposure assessment 260

9.6.4 Method for controlling biogenic amines in food 263

9.7 Modelling of growth and inactivation: kinetic approaches 264

9.7.1 Model categories: an overview 264

9.7.2 Growth - no growth interface: a probabilistic approach 267

9.7.3 Modelling of toxin production 268

9.8 Conclusions 268

References 269

10 Regulatory Measures for Microbial Toxins 287
M. Hickey

10.1 Introduction and background 287

10.2 The evolution and economic significance of heat-treated milk and milk products 288

10.2.1 Fluid milk 289

10.2.2 Evaporated and sweetened condensed milks 289

10.2.3 Milk and dairy powders 290

10.2.4 Cheeses (natural and processed) and fermented milks 291

10.3 Bacterial toxins 292

10.3.1 Staphylococcal enterotoxins 292

10.3.2 Bacillus cereus toxins 293

10.3.3 Clostridium botulinum toxins 294

10.4 Regulatory provisions on bacterial toxins in milk and milk products 297

10.4.1 European regulations on food hygiene and food safety 297

10.4.2 US milk hygiene and food safety standards 303

10.4.3 International perspective on food hygiene and safety – Codex Alimentarius 307

10.5 Mycotoxins 310

10.5.1 Aflatoxins 311

10.5.2 Other mycotoxins 311

10.5.3 Aflatoxins M1 and B1 and their regulatory provisions 312

10.5.4 EU legislations on aflatoxins in milk, milk products and animal feed 313

10.5.5 Regulations of aflatoxins of importance in milk and milk products in the USA 314

10.5.6 Regulations of aflatoxins of importance in milk and milk products in Canada 314

10.5.7 Regulation of aflatoxins in Australia and New Zealand 315

10.5.8 MERCOSUR standard on aflatoxins 315

10.6 Conclusions 316

References 316

Index 323

Adnan Y. Tamime is a Consultant in Dairy Science and Technology, Ayr, UK. He is the Series Editor of the SDT's Technical Book Series.