Executive Summary

In 1992 a working group of the UK Advisory Committee on the Microbiological Safety of Food presented a report on Vacuum Packaging and Associated Processes regarding the microbiological safety of chilled foods. The report supported subsequent guidance provided by the UK Food Standards Agency for the safe manufacture of vacuum packed and modified atmosphere packed chilled foods. In 2021 the ACMSF requested that a new subgroup should update and build on the 1992 report as well as considering, in addition to chilled foods, some foods that are intended to be stored at ambient temperatures. The new subgroup agreed a scope that includes the conditions that support growth and/or neurotoxin formation by C. botulinum, and other clostridia, as well as identification of limiting conditions that provide control. Other foodborne pathogens that need to be considered separately and some foods including raw beef, pork and lamb were explicitly excluded.

The subgroup considered the taxonomy, detection, epidemiology, occurrence, growth, survival and risks associated with C. botulinum and other neurotoxin-forming clostridia. There has been no significant change in the nature of foodborne botulism in recent decades except for the identification of rare cases caused by neurotoxigenic C. butyricum, C. baratii and C. sporogenes. Currently evidence indicates that non-clostridia do not pose a risk in relation to foodborne botulism.

The subgroup has compiled lists of incidents and outbreaks of botulism, reported in the UK and worldwide, and have reviewed published information concerning growth parameters and control factors in relation to proteolytic C. botulinum, non-proteolytic C. botulinum and the other neurotoxigenic clostridia.

The subgroup concluded that the frequency of occurrence of foodborne botulism is very low (very rare but cannot be excluded) with high severity (severe illness: causing life threatening or substantial sequelae or long-term illness). Uncertainty associated with the assessment of the frequency of occurrence, and with the assessment of severity, of foodborne botulism is low (solid and complete data; strong evidence in multiple sources).  The vast majority of reported botulism outbreaks, for chilled or ambient stored foods, are identified with proteolytic C. botulinum and temperature abuse is the single most common cause. In the last 30 years, in the UK and worldwide where a cause can be identified, there is evidence that known controls, combined with the correct storage, would have prevented the reported incidents of foodborne botulism.

The subgroup recommends that foods should continue to be formulated to control C. botulinum, and other botulinum neurotoxin-producing clostridia, in accordance with the known factors. With regard to these controls, the subgroup recommends some changes to the FSA guidelines that reflect improved information about using combinations of controls, the z-value used to establish equivalent thermal processes and the variable efficacy associated with some controls such as herbs and spices. Current information does not facilitate revision of the current reference process, heating at 90°C for 10 minutes, but there is strong evidence that this provides a lethality that exceeds the target 6 orders of magnitude reduction in population size that is widely attributed to the process and the subgroup includes a recommendation that the FSA considers this issue.

Early detection and connection of cases and rapid, effective coordinated responses to very rare incidents are identified as crucial elements for reducing risks from foodborne botulism. The subgroup recommends that the FSA works closely with other agencies to establish clear and validated preparedness in relation to potential major incidents of foodborne botulism in the UK.

 

 

Terms of Reference

• Review the risk posed by botulinum neurotoxin-producing clostridia in foods stored at ≤ 8°C that support growth or toxin production.

• A preliminary assessment of the risk posed by botulinum neurotoxin-producing clostridia in food designed to be stored at ambient temperature that supports growth or toxin production.
• Where appropriate, consider other risk-related evidence relevant to neurotoxin-producing clostridia during the lifetime of the group.

 

Scope

Conditions that support growth and/or neurotoxin formation by C. botulinum and other clostridia. Where practical this includes the identification of a limiting condition that allows growth and/or neurotoxin formation by C. botulinum as well as identification of a limiting condition that provides control. Preliminary consideration of non-clostridia that have gene sequences homologous with the botulinum neurotoxin genes is included.

 

The following will be excluded from consideration by the committee, as a wealth of evidence is already in existence:

• Foods given a botulinum cook;
• Foods such as honey that are associated with infant botulism/intestinal colonisation;
• Vacuum packaged/modified atmosphere packaged raw beef, pork and lamb (without added ingredients), to the extent that they have already been assessed by a previous ACMSF subgroup;
• Listeria monocytogenes, Bacillus cereus and other pathogens not specified in the scope or the terms of reference that need to be considered separately.

 

 

Working Group List of Members

 

Chair

Dr Gary Barker

 

Members

Dr Wayne Anderson

Dr Roy Betts  

Mr Alec Kyriakides

Miss Heather Lawson

Prof Peter McClure

Prof David McDowell

Prof Mike Peck

 

Secretariat

Mr Adekunle Adeoye

Dr Paul Cook

Ms Victoria Cohen

Dr Iulia Gherman

 

FSA/FSS Representatives

Dr Karen Pearson

Mr Ian Woods

Acknowledgements

Information provided by:

Dr Kathleen Glass, University of Wisconsin-Madison

Dr Gauri Godbole, UK Health Security Agency & University College London

Karin Goodburn MBE, Chilled Foods Association

Dr Kristin Marie Schill, University of Wisconsin-Madison

Paul Thomas, Specialist Cheesemakers Association

Dr Arnoud van Vliet, University of Surrey

 

 

Glossary

Anaerobic broth. Anaerobic microbiological broth, a nutrient medium used to grow bacteria in laboratory settings as a substitute for foods.

Decimal reduction time (D-value), DT. The time required, at a fixed temperature T, for heat to reduce the size of a population by 90% (The logarithm of the D value is frequently used as an alternative expression).

Enzyme-Linked Immunosorbent Assay, ELISA. An investigative procedure to measure antibodies, antigens, proteins and glycoproteins in biological samples.

Mouse Lethal Bioassay, MLB. An animal test conducted to ascertain the presence/absence of botulinum neurotoxins.

pH. A scale of acidity from 0 to 14. More acidic solutions have lower pH and more alkaline solutions have a higher pH; neutrality has pH = 7.0.

Polymerase Chain Reaction, PCR.  A laboratory technique for rapidly producing billions of copies of a specific segment of DNA, which can then be studied in greater detail.

Whole Genome Sequencing, WGS. A comprehensive method for analysing entire genomes.

Multi-Locus Sequence Typing, MLST. An unambiguous procedure for characterising isolates of bacterial species using the sequences of internal fragments of (usually) seven house-keeping genes.

Hazard Analysis and Critical Control Points, HACCP. Principles on which a food safety management system should be based; that is a risk assessment for food production covering all aspects from supplier to customer.

Dark/Ghost kitchens. Physical kitchen premises that exist solely for the delivery market to provide food to the public. There is no dine in or collection option for customers.

z-value, z. The difference in temperature of two isothermal heating processes that have decimal reduction times that differ by a factor of 10 (some conventions express z-value in units ‘centigrade degrees’ to emphasise the significance of temperature difference).

Water activity, aw. The chemical potential for water molecules in a system that includes water, ionic species and dissolved solids; alternatively called water availability. Pure water has aw = 1.

Redox potential, Eh. Sometimes called the reduction oxidation potential, measured in millivolts (mV), it is the tendency of food to release electrons (low redox Eh = 100 to -300 mV) or accept electrons (high redox Eh = 300 – 500 mV).

Salt. In this report ‘salt’ refers specifically to sodium chloride or NaCl.

 

1. Introduction and Background

In 1992 a working group of the UK Advisory Committee on the Microbiological Safety of Food (ACMSF) presented a report on Vacuum Packaging and Associated Processes with regard to the microbiological safety of chilled foods¹. The report had particular emphasis on the risks associated with psychrotrophic (non-proteolytic) Clostridium botulinum in chilled foods. The report was widely acknowledged and supported subsequent guidance provided by the UK Food Standards Agency, and many other organisations, for the safe manufacture of vacuum packed and modified atmosphere packed chilled foods.

FSA guidance published in 2020² recommends “that, in addition to chill temperatures (3 _-8°C) which should be maintained throughout the food chain, the following controlling factors should be used singly or in combination to prevent growth and toxin production by non-proteolytic C. botulinum in chilled foods with a shelf-life of more than 10 days;

• a heat treatment of 90°C for 10 minutes or equivalent lethality at the slowest heating point in the food
• a pH of 5.0 or less throughout the food and throughout all components of complex foods
• a minimum salt level of 3.5% in the aqueous phase throughout the food and throughout all components of complex foods
• a water activity (aw) of 0.97 or less throughout the food and throughout all components of complex foods
• a combination of heat and preservative factors which can be shown consistently to prevent growth and toxin production by non-proteolytic C. botulinum”

 

In 2021 the ACMSF requested that a new subgroup should update and build on the 1992 report. The new subgroup was requested to review new knowledge concerning C. botulinum and the formation of botulinum neurotoxin and to consider, in addition to chilled foods, some foods that are intended to be stored at ambient temperatures. The terms of reference for the subgroup, and an indication of restrictions on the scope of the review, are included as a preface to this report.

Since the publication of the 1992 report a wealth of information on C. botulinum, botulinum neurotoxin-producing clostridia and on food processing and packaging technology has been published. The types of food now being produced, corresponding production and distribution technology and the way hazards and risks are assessed and managed have evolved. The 1992 report covered risks from C. botulinum growth and toxin formation in chilled foods and also addressed other pathogens of concern. In 1992, the ACMSF considered foods stored at temperatures of “10°C and below” but, soon after, regulation stipulated that chilled foods should be stored “at or below 8°C”. For clarity, in this report, chilled storage corresponds to temperatures that do not exceed 8°C.

As in 1992 microbiological food safety is the issue of primary concern for this report. Contributory considerations include:

• Further information has been published on the prevalence and risk associated with non-proteolytic C. botulinum in chilled foods and the impact of this information on the risk assessment conducted by the ACMSF in 1992 requires corresponding review.

• The developments in food processing and packaging technology that have occurred since the publication of the 1992 report require consideration from the perspective of microbiological food safety. New packaging techniques and materials have become more widely used for controlled atmosphere packing and there are movements towards longer shelf life to reduce food waste, lower heat treatments to reduce use of energy, reduced preservation with salt or nitrites and alternative food ingredients as part of a move away from meat that merit consideration from a food safety perspective.
• It has become clear that there are foods on the market that are designed to be stored in ambient conditions that could support the growth of, or toxin formation by, botulinum neurotoxin-forming clostridia. A risk assessment for these types of food is required; this will extend the scope of the consideration to cover clostridia that can only grow and produce toxin at temperatures above those considered to be chilled.
• In the 1992 report C. botulinum was considered to be the sole species responsible for the production of botulinum neurotoxin. Recent scientific evidence has shown that clostridial species other than C. botulinum contain genes responsible for the production of botulinum neurotoxin and some have been associated with outbreaks of botulism. Additionally, some bacteria of non-clostridial genera have been found to include gene sequences that have some similarity with the C. botulinum neurotoxin genes. As previous risk assessments in relation to foodborne botulism have focussed almost exclusively on C. botulinum, it is important to review the significance of the new findings in relation to the safety of food.

Some background for this consideration is included in further sections of this introduction and details relating to taxonomy, detection, epidemiology, microbiology and risks are included in subsequent chapters. This report concludes with some recommendations in relation to microbiological safety of foods.

1.1 Clostridia

The genus Clostridium is composed of a wide range of Gram-positive, spore forming, rod shaped bacteria. They are considered to be anaerobic although the strictness of a requirement for anaerobiosis has been reported to be quite varied between species and strains. The presence of high levels of oxygen does not, in isolation, guarantee food safety with respect to C. botulinum or other toxigenic clostridia. Details of factors controlling growth and survival are included in Chapter 5 of this report.

Clostridium species can be found in a wide range of different environments including in foods where some species can cause food spoilage and others food poisoning. Of the species that cause food poisoning, Clostridium perfringens and the botulinum neurotoxin-producing clostridia are the most important. C. perfringens is responsible for outbreaks of food poisoning and is often associated with meat and meat products. These organisms can sporulate in the small intestine producing large amounts of C. perfringens enterotoxin which causes illness³; this hazard is beyond the scope of this review.

1.2 C. botulinum

The illness caused by C. botulinum was recognised in the early nineteenth century but it was not until the 1890s that it was attributed to a toxin produced by a bacterium; the bacterium was initially named Bacillus botulinus.  Later investigations of outbreaks indicated that there were different types of causative organisms, some being proteolytic and others non-proteolytic, and serological studies indicated that different types of neurotoxins could be produced by different strains. Early in the 20^th century B. botulinus was categorised as belonging to the genus Clostridium in a classification that separated the aerobic members of the genus Bacillus from the anaerobic members of the genus Clostridium.

1.3 The botulinum neurotoxin-forming clostridia

Originally C. botulinum was considered the only species able to produce botulinum neurotoxin. However, as microbial species differentiation has improved, it has become clear that the situation is more complex and that the traditional view of the species and its various groups has changed. Details of taxonomy for botulinum neurotoxin-forming clostridia are included in Chapter 2 of this report.

The scope of the 1992 ACMSF report was limited to chilled foods so only the risks associated with non-proteolytic C. botulinum, and risks associated with other pathogens that can survive and grow at low temperatures, were considered. Consideration of foods stored at ambient temperatures means that risks associated with proteolytic C. botulinum are within the scope of this update. Details of the conditions that support growth and survival of C. botulinum are included in Chapter 5 of this report.  

1.4 Non-clostridial species

The development of genome sequencing techniques has resulted in the discovery of botulinum neurotoxin-like sequences within the genomes of some non-clostridial species. These have not been associated with botulism. This was first reported following the identification of a botulinum neurotoxin-like gene sequence in Weissella oryzae⁴. Further details are included in Chapter 2 of this report.

1.5 Botulinum neurotoxins

Although botulinum neurotoxins are now known to be produced by species other than C. botulinum, the broad group of toxins are still referred to as botulinum neurotoxins.  C. botulinum neurotoxins are a diverse range of proteins represented by at least seven serotypes and more than 40 subtypes⁵. New clostridial strains producing novel toxin variants are still being identified.  The toxins are proteins made up of a heavy chain and a light chain. The role of the heavy chain is reported to be binding of the toxin to receptors in the peripheral nerve and translocation of the light chain into the nerve cell cytoplasm. The light chain blocks the release of the neurotransmitter acetylcholine which leads to flaccid paralysis and botulism⁵. Toxin types A, B, E and occasionally F have predominantly been associated with human foodborne botulism. Toxin types C and D mainly cause disease in animals and have rarely been linked to human foodborne botulism. Toxin types G and X have not been associated with foodborne botulism.  Modern techniques have identified a range of hybrid toxins. Details of the taxonomy of botulinum neurotoxins are included in Chapter 2 of this report.

1.6 Botulism - the illness

The botulinum neurotoxins are the most potent naturally occurring toxins. Human foodborne botulism is a severe, and potentially lethal, neuroparalytic intoxication potentially caused by the consumption of as little as 50 ng of botulinum neurotoxin. The classic symptoms of botulism include a severe flaccid muscle paralysis. The WHO provides the following description of foodborne botulism⁶: “Early symptoms include marked fatigue, weakness and vertigo, usually followed by blurred vision, dry mouth and difficulty in swallowing and speaking. Vomiting, diarrhoea, constipation and abdominal swelling may also occur. The disease can progress to weakness in the neck and arms, after which the respiratory muscles and muscles of the lower body are affected. There is no fever and no loss of consciousness. Symptoms usually appear within 12 to 36 hours (within a minimum and maximum range of 4 hours to 8 days) after exposure. The disease can be fatal in 5 to 10% of cases”.

Human botulism tends to be categorised according to how the toxin enters the body (exposure) and several types have been identified:

• Foodborne botulism:  toxin is formed during growth of the organism in food and when the food is consumed the pre-formed toxin enters the body causing illness.
• Infant botulism: C. botulinum spores are consumed in food and then germinate within the gut before cells grow and form toxin in situ. This type of botulism is associated with infants under one year old.
• Wound botulism: spores of C. botulinum germinate before cells grow and produce toxin in a wound. This type of botulism has, in recent years, been associated with injected drug use.
• Adult infectious botulism: a rare type of botulism with causation similar to infant botulism.
• Iatrogenic botulism: illness is caused by an accidental overdose of therapeutic botulinum neurotoxin.
• Inhalation botulism: illness is caused by inhalation of aerosolised neurotoxin. 

1.7  Industrial and domestic practices

In 1992 contemporary food safety often involved extremes of freezing and heating, in industrial settings, for relatively unprocessed foods as well as fermentation, drying and acidification. Processing was generally followed by storage in sealed metal or robust plastic containers. The 1992 ACMSF report was in response to the increasing use of packaging technologies that were designed to improve the quality and extend the shelf life of chilled foods. Packaging technologies for chilled foods have continued to change over time with development of new packaging materials, use of skin packing, the movement towards recyclable materials and the push for longer shelf life (to increase production efficiency, meet the needs of modern food distribution chains and to reduce food waste). In addition it is clear that, since 1992, for ambient stable foods there have been some developments that do not include traditionally accepted controls, and could theoretically allow germination, growth and toxin formation by C. botulinum leading to foodborne botulism; vegetables stored in oil are one example. Reports of botulism indicate that a significant number of incidents originate from the preparation or preservation of food in non-industrial environments. Historically, notably outside the UK, domestic and small scale non-industrial procedures such as home canning, bottling, preservation in oil and home fermentation have all resulted in incidents of foodborne botulism. Details of worldwide botulism incidents are included in Chapter 4 of this report. In a majority of cases the root cause of domestic outbreaks of foodborne botulism is the absence, or incorrect use, of C. botulinum control measures.

Drivers and trends in the UK point to an expanding role for home preservation and small scale non-industrial supply of chilled and ambient stored foods that cannot be neglected as a possible emerging source of foodborne botulism. Evolving systems for off-site or ghost supply, for buying and selling food online, and in social media marketplaces have implications for food safety. With respect to foodborne botulism the potential extent of additional risk for UK consumers, where there is little tradition for home preservation etc., is unknown and the facilities for the application of uniform controls, or effective traceability, in non-commercial food supply chains are not apparent. It may be prudent to establish greater awareness of foodborne botulism amongst the growing population of emerging small scale food producers.

The name Clostridium botulinum has been used for bacteria that form botulinum neurotoxin. For many decades it has been recognised that C. botulinum is not a distinct species but a collection of diverse clostridia that share the common property of forming botulinum neurotoxin⁷. Genes potentially encoding a botulinum neurotoxin have also recently been detected outside of the genus Clostridium.

Since 1992 scientific developments, notably whole genome sequencing (WGS), have provided a better understanding of the taxonomy and diversity of botulinum neurotoxin-forming clostridia and their neurotoxins. Information from WGS is valuable when selecting strains for challenge test experiments and process validation, is a valuable resource for improved pathogen detection and discrimination, is vital for tracing outbreaks and is central to studies of pathogen biology and evolution for C. botulinum and its neurotoxin genes.

This new information and understanding indicates it is unlikely that properties of botulinum neurotoxin-forming clostridia and their neurotoxins, or the risk of human foodborne botulism that they present, have changed in recent decades (although changes in food processing may impact on risk).

The diverse collection of clostridia that form botulinum neurotoxin can be separated into six genomically and phenotypically distinct groups (Table 1). These six groups are sufficiently distinct for each to be considered a separate bacterial species. Two of these groups, proteolytic C. botulinum (including C. sporogenes) and non-proteolytic C. botulinum, are strongly associated with human foodborne botulism, with the other groups either weakly associated or not associated. Each group includes highly related strains that do and do not form botulinum neurotoxin. There has been a greater interest in sequencing strains that form botulinum neurotoxin rather than non-toxic strains. Thus, for each group, the fraction of sequenced toxic strains is likely an overestimate of the fraction of toxic strains found in the environment.

WGS has provided details of the taxonomy, population structure and diversity within each group, and evidence of horizontal transfer of botulinum neurotoxin genes between distantly related clostridia. The gene encoding the botulinum neurotoxin is located on the chromosome or a plasmid or other mobile genetic element.

Different naming conventions are currently used for botulinum neurotoxin-forming clostridia, and the present situation is dynamic. In this report the framework in Table 1 is used for clarity. Further details of the association with human foodborne botulism are given in Chapter 4 of this report.

 

 

Table 1

Six genomically and phenotypically distinct groups of botulinum neurotoxin-forming clostridia

Name of botulinum neurotoxin-forming clostridia used in this report	Association of botulinum neurotoxin-forming clostridia with human foodborne botulism	Serotypes of botulinum neurotoxin formed
Proteolytic C. botulinum/C. sporogenes	Strong	A, B, F, X
Non-proteolytic C. botulinum	Strong	B, E, F
C. botulinum Group III	Very weak	C, D
C. argentinense	No	G
C. butyricum	Weak	E
C. baratii	Weak	F

 

2.1 Properties of the botulinum neurotoxins

Eight antigenically distinct botulinum neurotoxin serotypes (A-G, X) have been identified in bacterial strains within six distinct clostridial groups (Table 1). The eight botulinum neurotoxin serotypes are separated into more than 40 botulinum neurotoxin sub-types⁵. Most sub-types have been identified through derivation of their amino acid sequence from the gene sequence and they differ by at least 2.6% in amino acid sequence from any other sub-type. Various hybrid neurotoxins have also been described. Human foodborne botulism is most frequently associated with botulinum neurotoxin serotypes A, B and E. While there is evidence that the botulinum neurotoxin serotypes show minor differences in potency, all botulinum neurotoxins must be considered to represent a major hazard.

2.2 Taxonomy of proteolytic C. botulinum/C. sporogenes group

The proteolytic C. botulinum/C. sporogenes group is frequently associated with human foodborne botulism. It comprises highly proteolytic mesophilic bacteria that form spores of high thermal resistance. WGS has separated this group into two major lineages that could be recognised as separate species⁸. Both lineages contain strains responsible for foodborne botulism, and include toxic and non-toxic strains. The first lineage contains strains of proteolytic C. botulinum, with most strains forming one or more botulinum neurotoxins of types A, B, and/or F (and in one case of infant botulism type X). These strains are strongly associated with foodborne botulism. The second lineage contains strains of C. sporogenes, with 20 out of 104 strains analysed in a recent study forming botulinum neurotoxin type B, and the remaining 84 strains non-toxic. The strains that form botulinum neurotoxin type B were distributed amongst closely related non-toxic strains, rather than clustering together. The toxic strains of C. sporogenes have been occasionally associated with foodborne botulism (including one case in the UK).

Recent findings include the recognition that the high thermal resistance strain PA3679 is a non-toxic strain of proteolytic C. botulinum rather than a non-toxic strain of C. sporogenes, and that some strains that have caused human botulism (foodborne, infant and wound) and form type B neurotoxin belong to C. sporogenes rather than to proteolytic C. botulinum⁸.

The identification of distinct lineages/clusters within the proteolytic C. botulinum/C. sporogenes group, and further genotypic and phenotypic characterisation of strains within these lineages/clusters has the potential to contribute to improved risk assessments. For example, strains within the proteolytic C. botulinum lineage seem to present a greater human foodborne botulism risk than strains within the C. sporogenes lineage, since (i) a greater proportion of strains form botulinum neurotoxin, and (ii) a greater fraction of strains has been associated with foodborne botulism. However, currently nothing prohibits other circumstances (e.g. alternative approaches to food processing) that might favour toxic strains in the C. sporogenes lineage.

Detailed information on the genotype and particularly the phenotype of botulinum neurotoxin-forming strains in the C. sporogenes lineage is presently limited. It is not known how closely the phenotype of neurotoxin-forming strains of proteolytic C. botulinum is followed compared to that of non-toxic strains of C. sporogenes. Further clarity on the homogeneity of the proteolytic C. botulinum/C. sporogenes group will be of great value in considering the human foodborne botulism risk, and in particular whether it may be appropriate to disaggregate their risks. Currently it is pragmatic to consider neurotoxigenic C. sporogenes together with proteolytic C. botulinum.

2.3 Taxonomy of non-proteolytic C. botulinum group

The non-proteolytic C. botulinum group is frequently associated with human foodborne botulism. It comprises psychrotrophic, weakly proteolytic bacteria that form spores of moderate thermal resistance. WGS has separated this group into two major lineages⁹. Both lineages contain strains responsible for human foodborne botulism. The first lineage includes most strains that form type E neurotoxin and also some non-toxic strains. These strains are frequently isolated from fish and the arctic/subarctic environment (including marine and freshwater). The second lineage is dominated by strains that form type B neurotoxin and also contains some strains that form type E or type F toxin and non-toxic strains. The strains are often isolated from European terrestrial environments (including pigs) and marine/freshwater environments.

Foodborne botulism outbreaks associated with strains within the type E toxin lineage frequently involve chilled fish products, while outbreaks associated with strains within the type B (dominant) toxin lineage often involve chilled meat products. This may reflect genotypic/phenotypic characteristics and/or the geographical location in which the strains are found. Preliminary data indicate that spores of strains in the type B (dominant) lineage may have higher thermal resistance than spores of strains within the type E toxin lineage¹⁰. The identification of distinct lineages within the non-proteolytic C. botulinum group, and the genotypic and phenotypic characterisation of neurotoxin-forming strains within these lineages will be of great value in evaluating the human foodborne botulism risk, and in particular whether it may be appropriate to consider risks from the two groups independently.

2.4 Taxonomy of C. botulinum Group III and C. argentinense

C. botulinum Group III has been only very weakly associated with human foodborne botulism (details of the associations are included in Chapter 4 of this report) and there is no evidence that it presents a new or increased risk. Strains within C. botulinum Group III are frequently associated with botulism in animals and birds. Clostridium argentinense has not been associated with human foodborne botulism. WGS has been used to describe strain diversity within these two clostridial groups, and strains possessing and lacking genes encoding botulinum neurotoxin appear closely related.

2.5 Taxonomy of C. butyricum group

The majority of strains of C. butyricum are non-toxic, but a fraction form botulinum neurotoxin type E and have been associated with human foodborne botulism on a few occasions. The WGS of a limited number of C. butyricum strains (67 strains including eleven toxic strains) are available in public databases, and although this dataset is very limited there is an indication that the toxic strains may form discrete lineages rather than being widely distributed amongst non-toxic strains¹¹. A detailed understanding of the homogeneity of the C. butyricum group will contribute to evaluating the human foodborne botulism risk presented by neurotoxigenic strains of C. butyricum.

2.6 Taxonomy of C. baratii group

The majority of strains of C. baratii are non-toxic, but a minority form botulinum neurotoxin type F and have been associated with human foodborne botulism on a few occasions. The WGS of only 16 strains of C. baratii (nine non-toxic strains and seven toxic strains) are presently available; including four toxic strains associated with a single outbreak of foodborne botulism. However, initial indications from this very limited dataset suggest that botulinum neurotoxin strains of C. baratii could be distributed amongst non-toxic strains^11–13. A thorough understanding of the genotypic and phenotypic homogeneity of the C. baratii group will contribute to evaluating the human foodborne botulism risk.

2.7 Non-clostridial strains that contain potential botulinum neurotoxin genes

Non-clostridial bacterial strains have not been associated with human foodborne botulism. Recent data mining of published WGS has led to the discovery of genes in non-clostridial bacteria with partial homology to genes that encode botulinum neurotoxins. These include Enterococcus faecium from a cow¹⁴, Weisella oryzae from fermented rice¹⁵ and Chryseobacterium piperi from sediment¹⁶. The putative neurotoxin sequences differed from those of the neurotoxin serotypes presently described in clostridia. Recombinant DNA methods indicate a potential for biological activity, although it is not yet established whether biologically active botulinum neurotoxins are formed by these bacteria.  It is presently unclear whether further non-clostridial strains contain undiscovered potential botulinum neurotoxin-encoding genes, and whether these strains could form botulinum neurotoxin, and potentially cause botulism. As some of these bacteria are from genera that are used as probiotics in humans and animals, a watching brief should be maintained on any future developments regarding neurotoxin gene carriage and expression.

2.8 Conclusion

The understanding of the taxonomy and diversity of botulinum neurotoxin-forming clostridia and their neurotoxins has improved over the last few decades and this progression is likely to continue. New information indicates that it is unlikely that the properties of botulinum neurotoxin-forming clostridia and their neurotoxins have evolved significantly since 1992. A detailed understanding of the genotypic and phenotypic homogeneity of the various groups of botulinum neurotoxin-forming clostridia will contribute to assessment of the human foodborne botulism risk that is associated with botulinum neurotoxin-forming strains within these groups.

Non-clostridial strains with potential botulinum neurotoxin-encoding genes have not been shown to form botulinum neurotoxin, and have not been associated with foodborne botulism, and based on present evidence should not be considered as food safety hazards. However, it is prudent to maintain a close watch on scientific reports that identify botulinum neurotoxin type gene sequences in non-clostridia and to develop clearly identified protocols for responding, rapidly, to any significant new information.

For neurotoxin-producing clostridia in food the detection process is multifaceted.  Detection refers to the detection and enumeration of spores (and/or viable organisms), to the detection and typing of toxin and to the detection and identification of toxin-encoding genes. Detection of neurotoxin in a laboratory setting, following a clinical diagnosis and subsequent treatment of illness, is paramount. In addition, the detection of toxin or spores of C. botulinum, in food or environmental samples, is a fundamental part of incident investigations and is a quantitative element contributing to scientific research programmes and risk assessments¹⁷. However, routine monitoring for the presence of spores, cells or toxin in food is not a practical component of food safety.

C. botulinum is a very serious human pathogen and detection involves potentially hazardous materials that require a high level of laboratory biosecurity at all stages. Laboratory work with C. botulinum is only pursued at a few specialist locations in the UK (including the UKHSA). Furthermore, because of the serious impacts associated with human botulism, the results of detection always require expert interpretation.

3.1 Healthcare settings

In emergency healthcare settings, a test to detect botulinum neurotoxin in a specimen of serum or faeces etc., immediately follows any clinical diagnosis of human botulism. Detection of toxin is not a requirement prior to the commencement of treatment and timely treatment with antitoxin is known to significantly reduce the severity of outcomes for human botulism cases. In an incident of foodborne botulism in the UK in 2011¹⁸ a clinical diagnosis followed two days after the initial report of symptoms. Neurotoxin was detected in a clinical sample, by Mouse Lethal Bioassay (MLB), approximately four days after the diagnosis and in a sample of food one day later (PCR assay identified spores and vegetative cells of C. botulinum in the food sample at approximately the same time as the MLB result).

The in vivo MLB is regarded as the gold standard for detection of botulinum neurotoxins in clinical and epidemiological settings. Results from the MLB establish, simultaneously, multiple elements of functionality for neurotoxin, including binding, internalisation and intracellular activity, which together confirm the ability to cause botulism. The MLB is effective for all toxin types and has sensitivity reported as 5-10 pg per ml. The MLB is usually performed with 0.5 ml intraperitoneal injection of an extract from the food or faecal sample; a human oral lethal dose of botulinum neurotoxin is estimated as ~1ng per kg bodyweight. In the clinical setting the MLB is routinely combined with an array of antitoxins (in cross neutralisation reactions) to provide information on toxin type, including complex situations that involve multiple toxin types, and strongly supports the tracking of outbreaks.

Although the MLB is very sensitive, robust and is strongly established, particularly from a public health perspective, it does have several drawbacks; not least a failure to meet the objectives of Reduction, Replacement and Refinement of animal tests (3 Rs) that are supported by many organisations including the European Union and the OECD.  A requirement for expensive specialist animal facilities, turnaround times ~4-5 days, and some issues surrounding the interpretation of negative results add to the ethical concerns and have stimulated exploration of improved alternative rapid and validated technologies for detection of botulinum neurotoxins¹⁹.

3.2 Technological solutions

Since 1992 detection for a range of toxin types and with sensitivity at least equal to that obtained by MLB has been achieved using a range of alternative methods²⁰.

• Modified in vivo, and ex vivo, methods dominantly employ stem cells or differentiated neurogenic cells to detect toxin activity directly. These methods are used mainly in research settings but do not improve on the slow turnaround time of the MLB and include a level of variability that is inconsistent with the demands of most clinical settings.
• Immunological detection systems, such as ELISA, that use antibodies specific to the botulinum neurotoxin antigens have rapid turnaround times (typically a few hours) and have been combined, effectively, with other techniques, such as mass spectrometry, to achieve good specificity. These methods can be automated, however, they may detect biologically inactive toxin.
• The enzymatic activity of botulinum neurotoxins (such as the ability to cleave the SNAP-25 protein) can be harnessed, often using a combination of specific antibodies, fluorescent dyes or mass spectrometry, to provide automated in vitro detection systems. The biological activity is strongly specific to each toxin type so that extension to detect a wide range of known types, and potential new toxins, is not trivial. In clinical settings the MLB is used to confirm biological activity in some scenarios.
• Nucleic acid-based methods^21,22, including PCR, real time PCR, MLST and WGS, can be used for the detection and characterisation of C. botulinum, botulinum neurotoxin-encoding genes, but not direct detection of toxin. Generally these methods are readily deployed in multiplex platforms but they do not establish biological activity. A large quantity of gene sequence information for C. botulinum is stored in easily accessible databases so that nucleic acid-based methods add powerful tools for use in epidemiological, surveillance and research settings.
• Multiplex PCR methods have been extended to include detection of genes that code for neurotoxins in C. butyricum and C. baratii.

Some of these methods have obtained regulatory approval but currently none satisfy the stringent requirements which are necessary to replace the MLB for the detection of botulinum neurotoxins in clinical settings.

3.3 Epidemiology settings

Epidemiological investigations are an essential response to human botulism outbreaks. Investigations include a search for toxins, or spores, in foods and food environments and prioritise detection methods that minimise false negatives, have rapid turnover and high throughput to maximise the potential for source tracking and risk management. In vitro immunoassays that can work with complex food matrices, sometimes used in combination with confirmatory MLB, are cost effective, operate well over a range of toxin concentrations and are used effectively to detect neurotoxin in public health investigations. Multiplex PCR assays have been developed to provide accessible, rapid and cost-effective tools for detecting and screening unknown isolates from environmental samples for genes encoding botulinum neurotoxins (but not necessarily corresponding to toxin-producing capability). PCR assays and WGS can provide additional phylogenetic information that has particular value in outbreak investigations.

In epidemiological settings qualitative detection of neurotoxins, or C. botulinum spores, is usually combined with characterisations of toxin type, or genotype, to facilitate source tracking (linking clinical cases with contaminated foods).  Increasingly the characterisation step uses sophisticated nucleic technology, with an appropriate amplification step, to establish type matches with high precision.

3.4 Detection of spores and cells

There are no selective culture media for C. botulinum so that the detection of spores or vegetative cells involves specialist microbiology. Detection of spores usually involves anaerobic incubation of heated food followed by the detection of toxin (or encoding gene) in the enrichment fluid. ELISA methods with specific sets of antibodies, or multiplex PCR techniques with specialised sets of primers, are used to confirm the detection of spores with toxin types A, B, E and F. Enumeration of spores is achieved by statistical analysis of detection in diluted samples. In research settings this method can detect a few spores per kilogram in food materials²³.

Spores, inactivated toxins, toxin antigens or clostridial DNA do not pose foodborne botulism risks directly (infant botulism, adult infectious botulism and wound botulism are exceptions) so that pre-emptive testing of materials for C. botulinum has limited value for ensuring food safety. However, reliable detection of toxin, spores or viable cells is an essential element of challenge tests that are a pillar of assurance during food product developments that involve new formulation or new processes.

C. botulinum and most clostridia reduce sulphite to sulphide under anaerobic conditions and the reaction is easily detected. However, this phenotype is common among many food bacteria, including many food spoilage bacteria and some non-spore forming bacteria, so that detection has minimal relevance for food safety (historically this reaction has been used as an indicator of hygiene for food production; particularly in the dairy sector). Genetic characterisations of sulphite-reducing spore formers, and sulphite reducing bacteria, indicate the possibilities for the development of a molecular assay, based on sulphite reduction genes, that has increased sensitivity²⁴.

3.5 Research settings

In research settings the detection and isolation of C. botulinum in food materials has helped to inform risk assessments, produced detailed maps of phylogenetic relatedness to support epidemiology and built a picture of diversity, novel toxins, genotypes and horizontal gene transfer possibilities that improves understanding, helps development of therapeutics and aids public health preparedness. Many research activities highlight ongoing searches for sequence specific primers and for high quality, pure, monoclonal antibodies to assist in the development of detection technologies for C. botulinum.

In silico detection methods, involving very powerful search techniques across multiple databases, have detected C. botulinum neurotoxin sequence homologs in a small number of non-clostridial strains but the biological significance is not, currently, understood.

3.6 Conclusions

Methods for the detection of botulinum neurotoxin in tissues and food materials are well established and provide strong support for clinical, epidemiological and research activities. Technological advances point to emerging test methodologies that maintain sensitivity and can potentially reduce the burden on the use of experimental animals and these developments should be widely supported.

Specialist laboratory techniques effectively detect spores, and viable cells, of C. botulinum in food materials and support research activity but do not establish surveillance as a realistic option for improved food safety.

 

At the time of the 1992 report, foodborne, infant and wound botulism were reported worldwide but foodborne botulism was by far the most common form reported in the UK. In contrast, currently in the UK, wound and infant botulism are more common forms than that associated with food.

There has been no significant change in the disease symptoms reported since 1992^25–27. It remains the case that foodborne botulism is such a rare disease in the UK that misdiagnosis is a possibility due to the association of symptoms, by General Practitioners, with other more common diseases^27–29. Misdiagnosis may result in delayed response to cases and outbreaks and may warrant increased awareness amongst the medical profession. The US Centers for Disease Control and Prevention (CDC) has issued clinical guidelines for the diagnosis and treatment of botulism²⁷.

Investigations into many outbreaks of botulism have focused on the determination of the toxin type, rather than on the identification of the organism and its phenotype, and this has led to the loss of valuable data for subsequent risk assessment.

In this chapter, cases and outbreaks of botulism are reviewed firstly with respect to their geography and then with respect to the type of organism and its associations and therefore some repetition of events is evident.

4.1 Botulism – the global picture

Botulism is regularly reported throughout the world and Table 2 summarises the cases of all types of botulism from selected countries where data are available. Foodborne botulism is responsible for most reports in many countries, but it is not universally the most prevalent form of botulism.  For example, in the USA, infant botulism is the most common form of botulism with 1862 reported cases between 2001 and 2017, vastly exceeding 326 foodborne cases. In the UK wound botulism causes significantly more cases than infant and foodborne botulism combined but this may partly reflect the nature of clinical investigations and their capability.

4.1.1 UK

The UK has an extremely low number of outbreaks of foodborne botulism with only 10 reported outbreaks involving 13 cases since 1992 (data to 2019⁸ are included in Table 3). There has been no increase in foodborne botulism in the UK in recent decades. There has been a noticeable change in the origin of foods associated with outbreaks; eight of the ten most recent outbreaks involved foods produced or illness acquired abroad. This highlights effectiveness of the safety controls applied by the UK food industry in relation to this organism. Food produced in the home caused the greatest number of outbreaks since 1992 and five of six outbreaks, where details of food storage were known, involved ambient storage. A single outbreak implicating chilled food was caused by temperature abuse. Where the cause or potential cause of an outbreak was identified the evidence indicates that established controls for C. botulinum, if applied correctly, would have prevented the incident.

4.1.2 Worldwide

A selection of outbreaks of foodborne botulism that have occurred globally since 1992 are detailed in Table 4³⁰. There has been no significant change in the nature of foodborne botulism in recent decades with the exception of the identification of rare cases caused by neurotoxigenic C. butyricum and C. baratii. A few outbreak strains originally thought to be proteolytic C. botulinum are now recognised as C. sporogenes (details are included later in this chapter). The proportion of botulism outbreaks implicating commercial foods is weighted towards chilled foods rather than ambient foods (details are included later in this chapter and in Table 4) and this may constitute a trend in relation to the food types causing outbreaks. However, it is possible that bias may have been introduced by supplementing the literature sources with review papers specifically on chilled foods. 42 of the 90 outbreaks reported involved non-commercially produced foods i.e. home produced, 40 involved commercially produced food and 8 were unclear. Temperature abuse was a contributory factor in 30 of 36 outbreaks where a cause was known or suspected. For commercially produced chilled foods (and one frozen food) where the cause was known or suspected 23 of 24 products were subject to some form of temperature abuse prior to the outbreak; the remaining product was consumed 3 days beyond the “Use By” date. Outbreaks implicating commercially produced products that were destined for ambient storage and where a cause was known or suspected occurred due to a variety of control failures including inadequate sterilisation, post-process contamination, inadequate formulation to prevent growth and toxin production and/or temperature abuse. In the case of both chilled and ambient storage of commercially produced foods, the established controls for C. botulinum, if appropriately applied, would have prevented these outbreaks. Novel food technologies are not a feature of reported botulism outbreaks which tend to implicate traditional technologies which have been widely employed for food production over many years e.g. canning.

4.1.2.1 France

France reported 402 outbreaks of human botulism between 1987 – 2016 consisting of 731 cases and 9 deaths³¹. Cooked ham from home-made preparation or from small scale producers was involved in 73.5% of the botulism outbreaks where the food source was identified. These outbreaks were mostly type B botulism; more specifically B4 (and non-proteolytic C. botulinum). The other sources of botulism were home-made canned vegetables or fruits (beans, asparagus, eggplant, spinach, pumpkin, chestnut), home-made meat or fish preparations and a small number of industrial foods (fish soup, chicken/beef sausage, chicken/enchiladas, ground meat, olives/dried tomatoes, fresh pasta carbonara). Most of the outbreaks with non-pork meat were type A botulism (and proteolytic C. botulinum). Two outbreaks of rare C. baratii neurotoxin type F (F7) botulism were observed in 2014 and 2015. Industrial ground meat prepared in a restaurant was identified as the source of one outbreak³². Previous cases of botulism attributed to C. baratii were generally associated with intestinal colonisation i.e. infant botulism or due to other predisposing conditions³³. The origin of the other C. baratii type F outbreak was not identified. Clostridium butyricum was implicated in a case of botulism in a 10-year-old boy with history of Meckel’s diverticulum and chronic constipation, presenting dysarthria, dry mouth, hypotony, respiratory failure, and cardiac arrest in 2011. Stool analyses were positive for C. butyricum neurotoxin E5 by PCR and DNA sequencing up to 2 months after discharge. Botulism by intestinal colonisation with neurotoxigenic C. butyricum from undetermined origin was strongly suspected³⁴.

4.1.2.2 Italy

Italy has one of the highest numbers of foodborne botulism cases in Europe with 1173 suspected foodborne cases between 1986 – 2015, of which 421 were laboratory-confirmed²⁵. Homemade canned foods were implicated in 80.5% (95/118 incidents, involving 143 persons) of confirmed outbreaks including an outbreak caused by restaurant canned green olives. Vegetables canned in oil and in brine/water were associated with 43.2% and 28.8%, respectively, of laboratory-confirmed outbreaks. Other types of food implicated in confirmed outbreaks were home-bottled tuna (7.6%), ham (5.9%), home-bottled meat (5.9%), salami/sausages (4.2%), cheese (2.5%) and tofu and seitan (1.7%). Among vegetables, the most frequent products involved in cases or outbreaks were mushrooms in oil, olives and turnip tops. Regarding fish products, home-canned tuna was the most common food linked to confirmed incidents. Cheese or dairy products were seldom associated with confirmed incidents despite a large outbreak caused by mascarpone cheese³⁵. Although not reported for foodborne botulism alone, 96% (316/330) of the laboratory confirmed incidents were due to toxin produced by proteolytic C. botulinum (neurotoxins Type B 79.1%, Type A 9.7%, Type F 0.3%, Type Ab 1.5% and Type Bf 0.6%). Of 36 cases of infant botulism in Italy from 1986 to 2015, C. butyricum neurotoxin type E was implicated in three cases. Botulism involving intestinal colonisation with C. butyricum neurotoxin type E was also found in two boys having a Meckel’s diverticulum³⁶.

4.1.2.3 USA

There were 326 laboratory confirmed foodborne botulism cases in the USA between 2001 and 2017 of which 277 implicated a food or beverage (a food or beverage was a laboratory confirmed source of botulinum neurotoxin in 156 cases)³⁷. 47% of cases implicated food prepared in the home excluding canned foods, 29% home canned foods, 10% commercially canned foods and 6% other commercially prepared foods (no food preparation method was available for 8% of cases). Neurotoxin types A, E and B were responsible for most of the cases (65%, 25% and 7%, respectively). Outbreaks implicating commercial foods included a chilli meal, chilli sauce and nacho cheese sauce. A case of botulism implicating spaghetti with sauce/meat was attributed to C. baratii type F³⁸.

4.1.2.4 Turkey

A systematic review of botulism cases in Turkey from 1983 to 2017 identified 91 foodborne cases³⁹. Not all cases or suspected foods were tested for botulinum neurotoxin but 10 of 19 tested were positive for type A toxin (proteolytic C. botulinum). The top-ranking food responsible for cases was canned green beans (30% and 28 reports). Other reported foods include strained yoghurt (x10), home-made local cheese (x24), canned purslane (x16), non-specific canned food, canned ferula communis or fennel (x4), canned peppers, scrambled eggs with garlic sausages, canned fried mushrooms (x5) and unknown.

4.1.2.5 Iran

A review of botulism cases in Iran in the period 2007 – 2017 identified 252 confirmed cases of foodborne botulism, 743 suspected foodborne cases and 48 fatalities⁴⁰. The most commonly implicated foods, accounting for 34.1% of events, were home-prepared traditional processed fish (smoked fish, salted fish, ham, bacon, blood pudding, mosaic salami and sausage). Other implicated foods were commercially canned fish (28.6%), fish spawn (10.5%), dairy products (10.1%), vegetables and home-prepared legumes (9.7%), cottage cheese (5.9%) and canned fruits (1.1%).

4.1.2.6 Canada

There were 91 laboratory-confirmed outbreaks of foodborne botulism in Canada between 1985 and 2005 (205 cases and 11 deaths). Seventy-five outbreaks were associated with non-proteolytic C. botulinum type E; seven outbreaks associated with type A neurotoxin and five outbreaks with type B neurotoxin. The non-proteolytic C. botulinum neurotoxin type E outbreaks were attributed to consumption of traditionally prepared marine mammal and fish products by native communities (principally the Inuit of Nunavik in northern Quebec and the First Nations population of the Pacific coast of British Columbia). Two botulism outbreaks were attributed to commercial ready-to-eat meat products (pâté and cooked boneless pork) and three outbreaks to foods served in restaurants (chopped garlic in oil, bottled chanterelle mushrooms and baked potato). All involved type A or type B toxin. A further eight outbreaks were attributed to non-native home-prepared foods⁴¹. A review of foodborne outbreaks in British Columbia in the period 2009 – 2013 identified 3 botulism outbreaks and 1 death⁴². Implicated foods include fruit and vegetables, seafood and sauces/condiments.

4.1.2.7 Poland

Reviews identified that 54 of 109 botulism cases in Poland between 2014 and 2017 were due to home-produced foods and 55 due to commercial foods. Canned foods accounted for the vast majority of cases (84) with canned meat (other than pork) being the most frequently implicated food followed by canned fish, sausages and cured meat, canned pork, canned meat and vegetables and canned mushrooms, fruits and vegetables^43–46.

4.1.2.8 China

A review of botulism in China between 2004 and 2020 identified a total of 80 foodborne outbreaks with 386 illnesses and 55 deaths²⁹. The most common foods implicated were home-prepared traditional processed stinky tofu and dried beef, accounting for 51.2% of outbreaks. Contributory factors causing the outbreaks included improper processing and improper storage (77.5% of outbreaks). Initial misdiagnosis of illness occurred in 27.5% of cases. In an overview of type E botulism⁴⁷, 11 outbreaks between 1965 and 2005 implicated soy bean milk, fermented bean curd, raw dried beef, dried mackerel and blood sausage. The bacteria involved were either non-proteolytic C. botulinum type E or C. butyricum type E.

4.2 Association of clostridia and non-clostridia with foodborne botulism

4.2.1 Proteolytic C. botulinum and C. sporogenes

The vast majority of botulism cases associated with foods are caused by proteolytic C. botulinum neurotoxin types A and B. Rare cases implicating foods have been reported involving toxin type F.

It is widely recognised that proteolytic C. botulinum and C. sporogenes are closely related species. Recent genomic studies have highlighted that a number of strains of proteolytic C. botulinum should actually be classed as strains of C. sporogenes and vice versa, and that neurotoxigenic and non-neurotoxigenic strains of proteolytic C. botulinum and C. sporogenes exist⁸ (details of Taxonomy are included in Chapter 2 of this report). Four outbreaks of foodborne botulism have been attributed to C. sporogenes type B, including one case in the UK, along with rare cases of infant and wound botulism. As the historical identification of proteolytic C. botulinum associated with cases or outbreaks of botulism was based on the neurotoxin type this does not strongly affect the data presented in this chapter; neurotoxigenic C. sporogenes would have been incorrectly assigned to proteolytic C. botulinum. Moving forward it may be necessary to redefine historical data on botulism.  

Proteolytic C. botulinum and neurotoxigenic C. sporogenes are far more resistant to adverse conditions than other C. botulinum groups, requiring stronger heat processes to destroy spores and lower pH and water activity to prevent growth in food. Foodborne outbreaks associated with proteolytic C. botulinum have not shown any significant change in pattern, in recent decades, that would indicate an increased risk to foods. The controls that have been in place to manage the survival, population growth and toxin production by proteolytic C. botulinum for many years appear robust and do not need modification (details of the Occurrence, Growth and Survival of C. botulinum are included in Chapter 5 of this report).

4.2.2 Non-proteolytic C. botulinum

Non-proteolytic C. botulinum neurotoxin types B and E (and very occasionally toxin type F) are associated with foodborne outbreaks of botulism, but generally to a lesser extent than proteolytic C. botulinum neurotoxin types A and B, although this does vary by country.

Non-proteolytic C. botulinum is much less resistant to processing used in the food industry than proteolytic C. botulinum. The spores are more readily destroyed by heat and growth is more readily controlled by pH or water activity than for proteolytic C. botulinum. However, non-proteolytic C. botulinum is able to grow at much lower temperatures than proteolytic C. botulinum; this includes growth at refrigeration temperatures (details of the Occurrence, Growth and Survival of C. botulinum are included in Chapter 5 of this report). This presents a risk in chilled, minimally processed, extended shelf-life foods and this led to the provision of FSA guidelines for manufacture and sale of chilled foods in the UK² (details of the FSA guidelines are included in Chapter 1 of this report).

Despite the risk presented by non-proteolytic C. botulinum in chilled foods there have been no reported outbreaks of botulism involving chilled foods from any commercial product where the food has been stored at the recommended chilled temperature and consumed within its designated shelf life. In a review of global botulism outbreaks⁴⁸ caused by chilled foods, in the period 1985 to 2015, 16 of 26 outbreaks identified (Table 5) were caused by proteolytic C. botulinum, one by C. baratii and four by non-proteolytic C. botulinum (all type E in vacuum packed fish). Five outbreaks did not identify the organism and three of these involved toxin type B (it is unknown whether proteolytic C. botulinum or non-proteolytic C. botulinum were causal).  Temperature abuse was identified as the cause or most likely cause in 25 of the 26 outbreaks and consumption of the product beyond its “Use By” date was the reported cause of the remaining outbreak. In a review of botulism outbreaks in the USA between 1994 and 2021 that involved commercially produced foods intended to be stored chilled, 11 events were due to unrefrigerated storage by the consumer at home and the other two were due to unrefrigerated storage by the retailer prior to sale⁴⁹. In three of the events refrigeration instructions were deemed to be inadequate and contributory to the outbreak. In the 10 events where a toxin type was determined, all involved proteolytic C. botulinum (nine type A and one type Bf).

The lack of evidence for outbreaks caused by non-proteolytic C. botulinum in correctly stored chilled foods in the UK, both before and after the introduction of recommendations following the 1992 report, together with a similar lack of evidence from other countries throughout the world, provides some useful context regarding the magnitude of the risk and consequent need for controls in chilled foods. However, this review has not extensively examined the potential for under-ascertainment of botulism cases and outbreaks in other countries nor the legislative or industrial controls applied to foods globally. Currently it is not possible to use this insight in revising the risk from non-proteolytic C. botulinum in vacuum and modified atmosphere packaged chilled foods. Further study of these factors may provide more conclusive evidence that could indicate a lower risk to chilled foods from this organism that might, in turn, merit reduced controls or a focus on foods where the risk is greatest due to the known frequency of contamination and the associations with outbreaks i.e. vacuum packaged fish and seafood.

4.2.3 C. botulinum Group III

C. botulinum Group III (toxin types C and D) is often associated with animal botulism but has been implicated in human disease on rare occasions. In a recent review of all nine of the type C and D botulism outbreaks cited in literature, eight implicated foods; one was associated with infant botulism²⁶. Of the food outbreaks five had food vehicles suspected or confirmed as pâté (x2), smoked chicken, diseased chicken and home-made ham. The vast majority (seven) of these outbreaks were reported before 1992 when methods of identification and typing were less well advanced. Sporadic increases in animal botulism especially in farmed animals such as cows, cattle and chicken have raised concern regarding the potential for increased risk of transmission to humans. However, the ACMSF has reviewed such matters on a number of occasions and has considered the risk to humans to be low and supported the adequacy of current controls^50,51. There is no evidence that C. botulinum Group III presents any new or increased risk in relation to human foodborne botulism.

4.3 Other neurotoxin-producing clostridia

 

4.3.1 Clostridium butyricum

Neurotoxigenic C. butyricum type E has been reported in a small number of cases of infant botulism³³, adult intestinal botulism³⁶ and foodborne botulism in India⁵²,  China⁵³  and Italy⁵⁴. In the Indian incident clinical samples were not analysed and, despite the organism being isolated from a crisp made from gram flour, it was not possible to definitively confirm foodborne botulism. In a Chinese outbreak that occurred in 1994 implicating salted and fermented paste made of soybeans and wax gourds⁵⁵ six cases were clinically diagnosed with neurotoxin type E botulism that was also confirmed in the food although C. butyricum type E was only isolated from the food following further studies several years later⁵³. Retrospective analysis has also indicated an association of C. butyricum type E with other historical outbreaks of foodborne botulism in China^47,56. Type E botulism in China is most commonly associated with fermented grain/beans and raw meat and is frequently reported far from the sea (e.g. in Qinghai-Tibet plateau at an altitude of approximately 4-5 km). Canederli (bread dumplings) were the suspected food in a case of foodborne botulism in Italy in 1999 involving C. butyricum neurotoxin type E. Neurotoxigenic C. butyricum type E is a relatively newly identified hazard. No foodborne outbreaks of this type have been reported in the UK. Low numbers of confirmed outbreaks in other countries indicates that the risk presented by this organism has not substantially changed.

4.3.2 Clostridium baratii

Clostridium baratii producing botulinum neurotoxin type F has been associated with several foodborne outbreaks of illness although it was first isolated in an infant botulism case in the USA⁵⁷ and has subsequently been associated with a number of further cases of infant botulism^38,58 and adult intestinal botulism^59,60. Foodborne outbreaks have been reported in Spain, implicating individual meat pit pies⁶¹ and in France, where the food source was not identified although the only common item consumed by the two cases was an alcopop (which tested negative for growth and toxin⁶²) that reportedly contained 5% alcohol and had pH = 3.5. An outbreak in France implicated frozen and defrosted ground beef used for the production of spaghetti Bolognese at a restaurant where temperature abuse was again suspected as the underlying cause^32,63. Spaghetti and sauce mixture was implicated in a case of botulism caused by C. baratii type F in the USA in 2001³⁸. Raw deer meat was associated with an outbreak of foodborne botulism in Thailand in 2006, and a strain of C. baratii type F was isolated and its genome sequenced^64,65.

Neurotoxigenic C. baratii is a relatively newly identified hazard. No foodborne outbreaks of this type have been reported in the UK. Low numbers of confirmed outbreaks in other countries indicates that the risk presented by this organism has not substantially changed.

4.3.3 Clostridium argentinense

Clostridium argentinense was originally isolated from soil samples in Argentina in the 1960s and despite being capable of producing botulinum neurotoxin type G, there are no cases of botulism that have been associated with this bacterium⁶⁵. C. argentinense is not covered further in this report.

4.3.4 Other bacteria with potential botulinum neurotoxin genes

The advent of genomics has allowed the identification of potential botulinum neurotoxin genes in a number of bacteria outside of the Clostridium genus (more details are included in Chapter 2 of this report). The presence of botulinum neurotoxin genes is of potential concern but none of these bacteria have been shown to be capable of forming botulinum neurotoxin in foods and there are no known cases or outbreaks of botulism associated with bacteria outside the genus Clostridium.

4.4 Conclusions

4.4.1 Outbreak identification and clinical diagnosis

Foodborne botulism in the UK is extremely rare but consequently this may result in delayed responses due to unfamiliarity in the diagnosis of cases and delays in reporting.

4.4.2 Organisms responsible for botulism

There has been no significant change in the nature of foodborne botulism in recent decades with the exception of the identification of rare cases caused by neurotoxigenic C. butyricum, C. baratii and C. sporogenes. Other clostridia and non-clostridia have been identified with botulinum neurotoxin genes but have not been implicated in cases of foodborne botulism in the UK or elsewhere.

4.4.3 Foodborne botulism trends

There has been a noticeable change in the nature of foods that are associated with outbreaks of foodborne botulism in the UK. Eight of the ten outbreaks in the last 30 years involved foods produced or illness acquired abroad and the majority of these involved home-production.

Botulism outbreaks implicating commercially produced chilled foods appear more prevalent than in previous decades although bias in ascertainment of evidence may have affected this conclusion. The vast majority of botulism outbreaks, for both chilled or ambient stored foods, are identified with proteolytic C. botulinum, i.e. those organisms that do not grow under chilled conditions, and temperature abuse is the single most common cause identified for foodborne outbreaks. In relation to outbreaks in the last 30 years, in the UK and worldwide where a cause can be identified, there is evidence to believe that known controls for the organism, combined with the correct storage of the foods, if applied correctly, would have prevented the incident. Novel food technologies are not a feature of botulism outbreaks and most implicate traditional technologies, such as canning, employed for food production.

4.4.4 Vacuum packaged, extended shelf-life chilled foods

There have been no reported outbreaks of botulism globally in chilled foods from any commercial product where the food has been stored at the recommended chilled temperature and consumed within its designated shelf life. There has not been an extensive review of the potential for under-ascertainment of botulism cases and outbreaks in other countries nor the legislative or industrial controls applied to foods globally. However, further study of these factors may provide evidence that could indicate a lower risk to chilled foods that might, in turn, merit reduced or more focused controls. Nevertheless, the evidence indicates that where chilled foods are associated with outbreaks of botulism this is almost exclusively due to temperature abuse and in most cases is caused by proteolytic C. botulinum.

 

Table 2

Botulism cases in selected countries

Country	Period	Foodborne cases	Infant botulism	Wound botulism (confirmed only)	Adult intestinal botulism	Inhalation	Iatrogenic	Other	Reference
UK	1992 - 2019	13 (A, B, Bf)	16 (A, B, Bf, E– C. butyricum)	112 (A, B, AB)	1 (NT)				Brunt et al., 20208
France	1992-2016	574 (A, B, AB, E, F - C. baratii)		1 (B)	1 (E – C. butyricum)	2 (B)			Rasetti-Escargueil et al., 202026
France	2004 - 2016		17 (A, B, AB, Bf)						Rasetti-Escargueil et al., 202026
Italy	1986 - 2015	421 (Not reported)	36 (A, B, Ab, Bf, E – C. butyricum)	6 (B)	3 (A, E – C. butyricum)				Anniballi et al., 201725
USA	2001 - 2017	326 (A, B, E, F)	1862 (A, B, Ab, Ba, Bf, E, F, F - C. baratii)	372 (A, B, AB)	10 (A, F)		7 (A, B)	5 F – C. baratii	Lúquez et al., 202137; CDC, 202266

Table 3

Summary of botulism cases recorded in the UK since the publication of the 1992 ACMSF Report on Vacuum Packaged Foods and underlying causes (Adapted from Brunt et al., 2020⁸).

Year	Cases	Organism (toxin type)a	Toxin subtype	Food	Country of origin	Storage (cause)	Reference
1998	2	Prot (B)	B2	Home produced bottled mushrooms	Italy	Ambient (Inadequate controlling factors)	McLauchlin et al., 200667
2003	1	Cspo (B)	B1	Home produced Sausage	Poland	Unknown (Inadequate controlling factors likely)	McLauchlin et al., 200667
2004	1	Un (un)	ntb	Commercial Hummus	UK	Chilled (Temperature abuse)	McLauchlin et al., 200667
2004	1	Prot (A)	nt	Unknown	Georgia	Unknown	McLauchlin et al., 200667
2005	1	NP (B)	B4	Home preserved pork	Poland	Ambient (Inadequate controlling factors)	McLauchlin et al., 200667
2010	1	Un (B)	nt	Unknown	Algeria	Unknown	Brunt et al., 20208
2011	3	Prot (A)	A1	Commercial korma sauce	UK	Ambient (Unknown cause)	Browning et al., 201118
2012	1	Prot (B)	B2	Commercial olives	Italy	Ambient (Unknown but inadequate controlling factors likely)	Brunt et al., 20208
2013	1	Un (un)	nt	Home produced Mushrooms	Poland	Ambient (Inadequate controlling factors)	Brunt et al., 20208; Brola et al 201368
2016	1	Un (B)	nt	Tuna??	Italy	Unknown storage and controlling factors	Brunt et al., 20208

^aProt: proteolytic C. botulinum, NP: non-proteolytic C. botulinum, Cspo: C. sporogenes, un, unknown.

^b not tested in the present study. 

 

Table 4

Reported outbreaks of botulism in foods occurring since the publication of the 1992 ACMSF Report on Vacuum Packaged Foods
(for UK, see Table 3).

Year	Location	Cases	Toxin type	Exposure source	Food vehicle	Laboratory-confirmed or suspecteda	Implicated/ suspected Food – typical storage	Cause	References
1990	United States	3	B	Point	Unclear	Confirmed	Surgeon fish (palani)	-	Kershaw, 199169
1991	Egypt	97	E	Point	Commercial	Confirmed	Uneviscerated gray mullet fish (faseikh) – not reported	Temperature abuse during processing b	Weber, 199370; Hibbs, 199671
1992	United States	4	E	Point	Non-commercial: other	Confirmed	Uneviscerated fish (moloha) – not reported	-	French, 199272
1993	United States	8	A	Intermittent common	Commercial	Confirmed	Cheese sauce - ambient	Contamination and temperature abuse b	Townes, 199673, Newkirk, 201274
1993	Italy	7	B	Intermittent common	Commercial	Suspected	Canned eggplant in oil - ambient	Inadequate controlling factors	D'Argenio, 199575
1994	United States	30	A	Intermittent common	Commercial	Confirmed	Baked potato dip - ambient	Temperature abuse	Angulo, 199876, Newkirk, 201274
1994	United States	1	A	Point	Commercial	Confirmed	Black bean dip - chilled	Temperature abuse - product not refrigerated	Sobel et al. 200477
1994	United States	2	A	Point	Commercial	Confirmed	Clam chowder - chilled	Temperature abuse - product not refrigerated	Sobel et al. 200477
1995	Canada	3	E	Point	Non-commercial: other	Confirmed	Fermented seal meat	-	Proulx, 199778
1995	Canada	5	E	Point	Non-commercial: other	Confirmed	Fermented walrus meat	-	Proulx, 199778
1995	Canada	2	B	Point	Commercial: other	Confirmed	Country style pâté– chilled	Temperature abuse – product not refrigerated	Leclair et al., 201341
1996	Italy	8	A	Intermittent common	Commercial	Confirmed	Mascarpone cheese – chilled	Temperature abuse	Aureli, 199679; Aureli, 200035
1996	India	34	Unknown	Point	Unclear	Suspected	Sevu (gram flour crisp)	-	Chaudhry, 199852
1996*	Spain	2	Unknown	Point	Non-commercial: other	Suspected	Green beans	-	Polo, 199680
1997	United States	1	Bf	Point	Commercial	Suspected	Burrito – chilled	Temperature abuse	Sobel et al., 200477
1997	Thailand	6	Unknown	Unclear	Non-commercial: home-canned	Suspected	Canned bamboo shoots	-	Swaddiwudhipong, 200081
1997	Germany	2	E	Point	Commercial	Confirmed	Hot-smoked whitefish VP – chilled	Unknown – Temperature abuse b	Korkeala, 199882
1998	Argentina	9	A	Intermittent common	Non-commercial: other	Suspected	Matambre meat roll	-	Villar, 199983
1998	Thailand	13	A	Point	Non-commercial: home-canned	Confirmed	Canned bamboo shoots	-	Pantukosit, 200784; Swaddiwudhipong, 200081; Wongwatcharapaiboon, 199985
1998*	United States	3	B	Point	Non-commercial: other	Confirmed	Peyote	-	Hashimoto, 199886
1999	Canada	4	B	Point	Non-commercial: home-canned	Suspected	Canned tomatoes	-	Loutfy, 200387
1999*	Turkey	4	Unknown	Point	Non-commercial: home-canned	Suspected	Uncooked canned vegetables	-	Erol, 199988
1999	France	1	A	Point	Commercial	Suspected	Fish soup – chilled	Temperature abuse – at home	Carlier et al., 200189
2000	France	9	B	Point	Non-commercial: home-canned	Suspected	Canned asparagus	-	Abgueguen, 200390
2001	United States	14	E	Point	Non-commercial: other	Suspected	Fermented beaver tail and paw	-	Horn, 200191
2001	United States	16	A	Intermittent common	Commercial	Confirmed	Frozen chili	Temperature abuse	Kalluri, 200392; Newkirk, 201274
2001	Iran	2	Unknown	Point	Unclear	Suspected	Salted fish	-	Vahdani, 200293
2001	Canada	2	E	Point	Non-commercial: home-canned	Confirmed	Jar of salmon roe	-	Dawar, 200294
2001	Canada	2	E	Point	Non-commercial: other	Confirmed	Stink eggs	-	Dawar, 200294
2001	Canada	1	A	Point	Commercial	Confirmed	Cooked boneless pork product - chilled	Temperature abuse – product not refrigerated	Leclair et al., 201341
2001	United States	1	F – C. baratii	Point	Non-commercial: other	Confirmed	Spaghetti noodles and meat sauce	-	Harvey et al., 200238
2002	United States	8	E	Intermittent common	Non-commercial: other	Confirmed	Raw beached whale (muktuk)	-	McLaughlin, 200495; Middaugh, 200396; Newkirk, 201274
2003	United States	1	A	Point	Commercial	Confirmed	Clam chowder	Temperature abuse	CDC, 200397
2004	United States	4	A	Point	Non-commercial: other	Suspected	Pruno	-	Vugia, 200998
2004	Germany	1	E	Point	Commercial	Suspected	Vacuum packed smoked salmon	Consumed 3d after Use By date	Dressler (2005)99
2004	Italy	28	B	Intermittent common	Commercial	Suspected	Green olives - ambient	Improper preservation	Cawthorne, 2005100
2004*	India	2	Unknown	Point	Non-commercial: other	Suspected	Canned meat, preserved curd	-	Agarwal, 2004101
2005	Turkey	10	A	Point	Non-commercial: other	Confirmed	Suzme yoghurt buried in soil	-	Akdeniz, 2007102
2005	United States	5	E	Intermittent common	Non-commercial: other	Confirmed	Salted fish	-	Sobel, 2007103
2005*	Turkey	5	Unknown	Point	Non-commercial: home-canned	Suspected	Canned mushrooms	-	Cengiz, 2006104
2006	United States, Canada	6	A	Intermittent common	Commercial	Confirmed	Carrot juice - refrigerated	Temperature abuse	Shuler, 2006105; Brown, 2010106; Sheth, 2008107
2006	Austria	5	Unknown	Point	Non-commercial: other	Suspected	Barbequed pork	-	Meusburger, 2006108; Topakian, 2009109
2006	Finland	2	E	Point	Commercial	Suspected	Smoked whitefish - chilled	Temperature abuse b	Lindstrom, 2006110
2006	United States	2	A	Point	Non-commercial: other	Confirmed	Fermented tofu	-	Meyers, 2007111
2006	Taiwan	5	B	Unclear	Non-commercial: other	Confirmed	Fermented raw goat meat (cinkrugan)	-	Tseng, 2009112
2006*	Italy	2	Unknown	Point	Non-commercial: other	Suspected	Preserved asparagus	-	Zanon, 2006113
2007	United States	8	A	Intermittent common	Commercial	Confirmed	Canned chili sauce - ambient	Processing failure	Juliao, 2013114; Newkirk, 201274; Ginsberg, 2007115
2007	China	66	A	Intermittent common	Commercial	Suspected	Sausage – not reported	Temperature abuse	Zhang, 2010116
2008	Turkey	8	B	Point	Commercial	Suspected	Unprocessed black olives - ambient	Unknown	Swaan, 2010117
2008	France	2	A	Point	Commerical	Confirmed	Chicken enchiladas - chilled	Temperature abuse – product not refrigerated and consumed 1 day after Use By date	King & the French Multidisciplinary Outbreak Investigation Team (2008)118
2008	Uganda	3	A	Point	Non-commercial: other	Suspected	Oil-based condiment	-	Viray, 2014119
2008	United States	6	A	Point	Non-commercial: home-canned	Confirmed	Canned carrots and green beans	-	Date, 2011120
2009	France	3	E	Point	Commercial	Suspected	Vacuum packed hot-smoked whitefish (VP) - chilled	Temperature abuse b	King, 2009121
2009	United States	3	A	Point	Non-commercial: home-canned	Confirmed	Canned green beans	-	Date, 2011120
2009	United States	3	A	Point	Non-commercial: home-canned	Confirmed	Canned asparagus	-	Date, 2011120
2010	France	5	A	Point	Unclear	Confirmed	Canned green beans or pork chops	-	Oriot, 2011122
2011	United States	8	A	Point	Non-commercial: other	Suspected	Pruno	-	Thurston, 2012123; Williams, 2014124
2011	United States	1	A	Point	Commercial	Suspected	Potato soup - chilled	Temperature abuse	CDC., 2011125
2011	United States	1	A	Point	Commercial	Suspected	Potato soup - chilled	Temperature abuse	CDC., 2011125
2011	Spain	5	F – C. baratii	Point	Unclear	Suspected	Indeterminate	-	Lafuente, 201361
2011	France	9	A	Intermittent common	Commercial	Confirmed	Green olive paste - ambient	Incorrect sterilisation	Pingeon, 2011126
2011	Finland	2	B	Intermittent common	Commercial	Confirmed	Jar of olives - ambient	Post process contamination	Jalava, 2011127; Forss, 2012128
2011*	Turkey	4	Unknown	Point	Non-commercial: home-canned	Suspected	Canned red peppers	-	Agacayak, 2011129
2012	Canada	3	E	Point	Commercial	Confirmed	Salt-cured fish (fesikh) – unknown need paper	Unknown – need access	Walton, 2014130
2012	United States	8	A	Point	Non-commercial: other	Confirmed	Pruno	-	Briggs, 2013131
2012	Japan	2	A	Point	Commercial	Confirmed	Adzuki batto (bean soup) (VP) – not reported	Temperature abuse b	Momose, 2014132
2012	United States	2	Unknown	Point	Commercial	Suspected	Broccoli soup - chilled	Temperature abuse	Edmunds et al., 202249
2012	Thailand	2	B	Point	Unclear	Confirmed	Fermented crab	-	Wangroongsarb, 2013133
2013	China	12	A	Point	Commercial	Confirmed	Smoked ribs (restaurant) – chilled	Temperature abuse b	Feng, 2015134
2013	Green- land	5	E	Unclear	Non-commercial: other	Suspected	Eider fowl	-	Hammer, 2015135
2013	Iran	5	Unknown	Unclear	Non-commercial: home-canned	Suspected	Canned cheese (kupeh)	-	Faridaalaee, 2013136
2014	France	2	F – C. baratii	Point	Commercial	Suspected	Alcopop?	Unknown	Castor, 201562
2014	Spain	3	Unknown	Point	Non-commercial: other	Confirmed	Stew	-	Buj-Jorda, 2015137
2014	USA	2	B	Intermittent common	Commercial	Confirmed	Jar of pesto - ambient	Unlicenced premise; inadequate processing	Burke, 2016138
2015	Slovakia	1	A	Intermittent common	Commercial	Confirmed	Hummus (pouches) Chilled	Temperature abuse b	Mad'arova, 2017139
2015	USA	29	A	Point	Non-commercial: home-canned	Confirmed	Canned potatoes	-	McCarty, 2015140
2015	France	3	F – C. baratii	Point	Non-commercial: other	Suspected	Meat (ground) frozen used in sauce by Restaurant	Temperature abuse	Trehard, 201663, Mazuet et al., 201712
2015*	New Zealand	1	Unknown	Point	Commercial	Suspected	Risotto - Chilled	Temperature abuse	Smyth et al., 2015141
2016	United States	1	A	Point	Commercial	Confirmed	Grain and vegetable product - chilled	Temperature abuse	CDC, 2016142
2017*	Spain	2	Unknown	Point	Commercial	Confirmed	Canned beans - ambient	Not reported	Ametller, 2017143
2017	USA	2	A	Point	Commercial	Suspected	Prepackaged Pouches of Liquid Herbal Tea -Chilled	Contamination and temperature abuse b	Kim, 2019144
2017	USA	10	A	Intermittent common	Commercial	Confirmed	Nacho cheese sauce (Pouch) - Ambient	Contamination and temperature abuse b	Rosen, 2020145
2018	Nigeria	3	Unknown	Point	Non-commercial: other	Suspected	Fish pepper soup	-	Okunromade, 2020146
2018	USA	3	A	Point	Non-commercial: home-canned	Confirmed	Canned peas	-	Bergeron, 2019147
2019	United States	4	A	Point	Commercial	Suspected	Potato product - chilled	Temperature abuse	Edmunds et al., 202249
2019	China	4	A, B, E	Point	Commercial	Suspected	Fish and ham (VP) – not reported	Not reported	Min, 2021148
2020	Italy	35	Unknown	Point	Unknown	Suspected	Salad	Unknown	FSN, 2020149
2020*	Portugal	1	Unknown	Point	Unknown	Suspected	Unknown	Unknown	Costa, 2021150
2021	United States	1	A	Point	Commercial	Confirmed	Clam chowder - chilled	Temperature abuse	Edmunds et al., 202249

^a Laboratory-confirmed or suspected food

^b Likely cause but not definitive

* Outbreak year not reported, publication year used instead

 

Table 5

Examples of foodborne botulism outbreaks involving commercial foods intended to be stored chilled (Peck et al., 2020⁴⁸).

Country (year)	Product	Organism (toxin type)	Cases (deaths)	Factors contributing to outbreak	References
Canada (1985)	Garlic-in-oil	Prot (B)	36	No preservatives; temperature abuse	St. Louis et al. (1988)151
UK (1989)	Hazelnut yoghurt	Prot (B)	27(1)	Toxin added with canned hazelnut conserve to correctly chilled yoghurt	O'Mahony et al. (1990)152
USA (1989)	Chopped garlic-in-oil	Prot (A)	3	Temperature abuse (product not refrigerated)	Morse et al. (1990)153
USA (1990)	Grilled fresh Palani (surgeon fish)	NR (B)	3	Temperature abuse	CDC (1991)154
USA (1993)	Canned cheese sauce (restaurant)	Prot (A)	8(1)	Contamination of canned cheese sauce after opening, then temperature abuse (opened tin not refrigerated)	Townes et al. (1996)73
USA (1994)	Potato dip (‘‘skordalia’’) and aubergine dip (‘‘meligianoslata’’) (restaurant)	Prot (A)	30	Toxin added with temperature-abused baked potatoes to correctly chilled yoghurt dishes	Angulo et al. (1998)76
USA (1994)	Clam chowder	Prot (A)	2	Temperature abuse (product not refrigerated)	Sobel et al. (2004)77
USA (1994)	Black bean dip	Prot (A)	1	Temperature abuse (product not refrigerated)	Sobel et al. (2004) 77
Canada (1995)	Country-style pâté	NR (B)	2	Temperature abuse (product not refrigerated)	Leclair et al. (2013)41
Italy (1996)	Mascarpone cheese	Prot (A)	8(1)	Temperature abuse; pH > 6	Aureli et al. (2000)35
Germany (1997)	Hot-smoked, vacuum-packed fish	NP (E)	2	Suspected temperature abuse	Korkeala et al. (1998)82
Argentina (1998)	Meat roll (“matambre”)	Prot (A)	9	Insufficient cooking, lack of preservatives, vacuum-packed in heat-shrunk plastic, & inadequate refrigeration	Villar et al. (1999)83
France (1999)	Fish soup	Prot (A)	1	Temperature abuse at home	Carlier et al. (2001)89
Canada (2001)	Cooked boneless pork product	Prot (A)	1	Temperature abuse (product not refrigerated)	Leclair et al. (2013)41
Germany (2004)	Vacuum-packed smoked salmon	NP (E)	1	Consumed 3 days after ‘Use By date’	Dressler (2005)99
UK (2004)	Organic hummus	NR	1	Time/temperature abuse	McLauchlin et al. (2006)67
Canada/USA (2006)	Refrigerated carrot juice	Prot (A)	6	Temperature abuse; product pH between 6 and 7	Sheth et al. (2008)107
Finland (2006)	Vacuum-packed smoked whitefish	NP (E)	1	Suspected temperature abuse	Lindström et al. (2006)110
China (2007)	Sausages	Prot (A)	66	Temperature abuse (product not refrigerated)	Zhang et al. (2010)116
France (2008)	Chicken enchiladas	Prot (A)	2	Time/temperature abuse (product not refrigerated); consumed 1 day after ‘Use By date’	King & the French Multidisciplinary Outbreak Investigation Team (2008)118
France (2009)	Vacuum packed hot-smoked whitefish	NP (E)	3	Suspected temperature abuse during travel and home storage	King et al. (2009)121
Italy (2010)	Cream of vegetable soup	NR (B)	1	Temperature abuse (product not refrigerated); long shelf-life	Daminelli et al. (2011)155
USA (2011)	Potato soup	Prot (A)	2	Temperature abuse (product not refrigerated)	CDC (2011)125
New Zealand (2015)	Chilled, ready-to-eat risotto	NR	1	Time/temperature abuse (product not refrigerated; consumed several months past best-before date)	Smyth et al. (2015)141
Slovakia (2015)	Hummus spread	Prot (A)	1	Suspected temperature abuse	Mad'arova et al. (2017)139
France (2015)	Frozen minced beef used in restaurant Bolognese sauce	C. baratii (F)	3	Time/temperature abuse (sauce prepared ≥24 h in advance, left at room temperature for several hours)	Mazuet et al. (2017) 32

Prot: proteolytic C. botulinum, NP: non-proteolytic C. botulinum, NR: not reported