Introduction

• About 900's: ¾Certain foods caused typical poisoning.

¾Emperor Leo VI of Byzantium forbade the manufacture of Clostridium botulinum blood sausages.

Introduction Introduction 1820 • 1793: An outbreak caused by blood sausages • Kerner collected was described in Wildbad, Germany data on 230 cases of typical poisoning. • Disease became known as "Kerner’s Disease."

Introduction Other Findings • van Ermengem isolated an anaerobic 1897 bacterium from cured raw ham that had caused “Kerner’s Disease” in 23 Extract from ham people and killed 3. & a culture of m.o.

killed a number of different experimental animals with the same signs as the disease in humans

1 Other Findings Introduction

• van Ermengem called the organism Bacillus botulinus after botulus. – Latin for sausage

• Later named C. botulinum proved to be fairly resistant

Introduction Back to Germany

• The C. botulinum strain isolated 1904 by van Ermengem was later Landman investigated botulism designated type B. caused by canned, white beans.

• The name of the disease was changed from Kerner’s Disease to botulism.

Landman Findings Cont.. How about the US?

1904 • The signs and symptoms were typical for botulism. • Between 1918-1922: ¾297 cases and 185 deaths • The antitoxin Landman produced did ¾mainly in California not cross-react with van Ermengem’s strain.

• Landman had discovered C. botulinum type A.

2 Introduction Introduction

• 1936: C. botulinum type E was isolated • 1922: C. botulinum type C was isolated ¾caused paralysis in chickens and cattle ¾from smoked fish that caused botulism in the US and Russia • 1929: C. botulinum type D was isolated ¾from cattle that died from paralysis

Introduction Introduction

• 1951: Wound botulism was described for the • 1960: C. botulinum type F was isolated in first time. Denmark ¾from liver paste that caused human botulism

Introduction Introduction

• 1985: Hall et al. found that a strain of C. • 1970: C. botulinum type G was isolated in barati produced type F botulinal toxin. Argentina • 1986: Aureli et al. and McCroskey et al. ¾from soil isolated strains of C. butyricum that ¾no reported cases of poisoning with this type produced type E botulinal toxin. in man or animals • 1973–1996: CDC documented 724 cases of verified foodborne botulism in American • 1976: Infant botulism was recognized. adults; mainly associated with home-canned vegetables.

3 Illness & Causative Agent Categories of Human Botulism • Botulism is a serious paralytic illness. 1. Foodborne botulism • It is caused by a nerve toxin that is produced by the bacterium. 2. Infant botulism • It is a rare illness. • It is much feared. 3. Wound botulism

4. Unclassified

Categories of Human Botulism Categories of Human Botulism • Foodborne botulism • Infant botulism ¾This type of food poisoning is ¾It was first recognized in 1976. caused by the ingestion of foods ¾This type of poisoning affects containing the potent infants under the age of 12 neurotoxin. months. ¾The neurotoxin is formed in ¾It is caused by the ingestion of C. the food during growth of C. botulinum spores. botulinum.

Categories of Human Botulism Categories of Human Botulism • Infant botulism ¾The spores germinate & • Infant botulism multiply, colonizing the ¾Infant botulism has been intestinal tracts of infants, and reported in 41 states nationwide. produce neurotoxin. ¾The incidence is 1 case per ¾The neurotoxin travels through the bloodstream to the central 100,000 live births. nervous system and causes ¾Case fatality rate is below 4%. flaccid paralysis.

4 Categories of Human Botulism Categories of Human Botulism • Infant botulism • Infant botulism ¾In California the incidence from 1985 to 1995 was 7.1 cases per ¾Honey is one vehicle that has 100,000 live births. been associated with infant ¾Estimated medical cost/case at botulism by a number of laboratory and $85,000 (total cost = $31 million). epidemiological studies.

Categories of Human Botulism Categories of Human Botulism • Wound botulism

• Infant botulism ¾This illness results from the pathogen itself infecting a wound. ¾Honey is now thought to ¾Foods are not the vehicle of account for no more than transmission. 5% of cases. ¾The microorganism produces the ¾California cases may come from spores on wind-blown neurotoxin which is transmitted to dust. other parts of the body via the blood. ¾Rare form of illness

Categories of Human Botulism Recorded Botulism Cases in the US: 1973–1996 • Unclassified Botulism type Range/yr Total (all ¾Resembles infant botulism, but affects years) adults. Food 8–86 724 ¾C. botulinum colonizes the intestinal tract of adults and produces the toxin Infant 0-99 1444 in vivo. Wound 0-25 103 ¾Thought to occur after antibiotic treatment depleted the indigenous Unclassified Not avail. 39 intestinal flora.

5 Classification of C. botulinum C. botulinum Toxins • There are seven types of C. botulinum • C. botulinum produces eight toxins (A, B, C , ¾A, B, C, D, E, F, and G 1 C2, D, E, F and G). ¾based on the serological • All are neurotoxins specificity of the neurotoxin except C . produced 2

Classification of C. botulinum C. botulinum Toxins • Types A, B, E, and, very rarely, F • Some strains produce pairs of are associated with human botulism toxins (foodborne, wound and infant • These are designated subtypes types). ¾The capital letter identifies the • Types Cand Daffect animals. type of toxin in greater amount • Type G has not been linked to ¾The lower case letter identifies illness up to this date. the type of toxin produced in lesser amount

C. botulinum Toxins Distribution of Serotypes in Human Botulism in the US • An example: Type Cases (%) Deaths (%) ¾strain isolated from a case of A 38 52 infant botulism was classified as subtype B 38 12 E 9.7 10 F 0 0 Unknown 13 0

6 C. botulinum Groups C. botulinum groups • C. botulinum strains are divided into • Another classification of C. botulinum four groups. strains is based on physiological ¾group I*: proteolytic and produce differences. neurotoxins type A, B, and F. ¾growth temperature ¾group II*: nonproteolytic and ¾pH produce neurotoxins type B, E, and F. ¾water activity *the most commonly involved in human ¾sodium chloride concentration illness.

C. botulinum Groups Characteristics of C. botulinum • C. botulinum strains are divided into four groups. • Gram positive ¾group III: variably nonproteolytic or • Sporeformer proteolytic and produce neurotoxins • Anaerobic type C and D. • Rods ¾group IV: proteolytic and produce • Produce a potent neurotoxin type G. neurotoxin

[A. Dowsett/Science Photo Library]

Characteristics of C. botulinum Characteristics of C. botulinum

• pH values for growth • Limiting water activity ¾Types A and proteolytic B (Gp I), ¾Type A 0.95 pH 4.6-8.5 ¾Minimum pH for E (Gp II) is: ¾Type B 0.94 ™ 6.2 at 5°C, and ™ 5.4 at 30°C ¾Type E 0.97

7 Characteristics of C. botulinum Characteristics of C. botulinum • Growth temperature • Limiting salt concentration for ¾Type A and proteolytic B (Gp I) growth 10–50°C ¾10.7–12% NaCl ¾E and non-proteolytic B and F (Gp II) 3.3–45°C ¾Non-proteolytic most sensitive ¾Spores are highly resistant to freezing

Characteristics of C. botulinum Characteristics of C. botulinum

• Redox potential • Heat resistance defined ¾Optimum growth occurs at ¾Decimal reduction time (D value; Eh of !350 mV 90% kill) ¾Time required to reduce the ¾E is the least anaerobic, microbial population by 1 log 0–100 mV cycle.

Characteristics of C. botulinum Characteristics of C. botulinum • Heat resistance • Heat resistance ¾121.1°C (250°F) / 3 min to ¾121.1°C (250°F): DRT = 0.20– achieve 1012-fold reduction 0.21 min for the most resistant (A (standard for low acid canned and proteolytic B) foods), “bot cook” ¾0.3–0.6 min causes 106-fold reduction and is standard for canned, cured meats

8 Characteristics of C. botulinum Mechanism of Toxin • Radiation resistance To cause 1012 fold reduction Neurotoxin 0 binds to neurons 0 ¾47 – 54 kGy for type A spores ¾10 – 11 kGy for type B spores internalized 0

¾7 – 9 kGy for type E spores prevents release of acetyl choline ¾12 kGy for type F spores (neurotrasmitter) ¾48 kGy is the accepted dose for sterilization of food spores

Nature of Food Botulism Clinical Symptoms • Intoxication • Symptoms include: • Onset is about 18 – 36 hrs after ¾nausea and vomiting ingestion of the food containing the neurotoxin. • Symptoms vary from a mild to severe illness.

Clinical Symptoms Nature of Illness

• Symptoms include: • Other symptoms include: ¾neurological signs ¾gastrointestinal problems ƒ blurred or double vision ƒ cramps ƒ difficulty in speaking or swallowing ƒ abdominal pain ƒ fatigue ƒ diarrhea, or ƒ lack of muscle coordination, and ƒ constipation ƒ difficulties in breathing

9 Pathogenic Dose • Few nanograms of C. botulinum Foods Implicated in Botulism neurotoxin can cause illness. • Any food that can support the growth • The neurotoxin produced is probably of this pathogen or allow the the most toxic compounds made by a biological system. germination of its spores and eventually toxin production can be • About 1 oz. (28.4 g) of this toxin can kill 200 million people. associated with this illness. • Fortunately, the incidence of the • Low acid foods (pH>4.6) illness is low.

Foods Implicated in Botulism Foods Implicated in Botulism • Home-canned or -preserved low-acid • North American Indian specialties vegetables ¾fish and fish eggs ¾asparagus, tomatoes, beans, ¾seal flippers mushrooms ¾peppers, corn, baked potato, chopped beets • Other implicated foods include luncheon meats, ham, sausage, ¾garlic in soybean oil smoked and salted fish, and lobster.

C. botulinum Outbreak C. botulinum Outbreak • In 1994, in Oklahoma, a 47-year old man was hospitalized for symptoms of • Upon testing: progressive dizziness, blurred vision, ¾The green beans tested negative for slurred speech difficulty swallowing, the toxin and nausea. • Twenty-four hours earlier the patient ¾The stew tested positive for the had eaten some home canned green toxin beans and beef and potato stew.

10 C. botulinum Outbreak Prevention • Assurance of destruction or • Apparently, inhibition of C. botulinum. ¾the stew was cooked, • Keep foods out of the ¾covered tightly, temperature danger zone (4.4 – 60°C or 40 – 140°F). ¾left out for four days at room • Botulinum toxin is destroyed by temperature, and heating at 80°C for 30 min or ¾then eaten without reheating. boiling or a few minutes. Thus re-heating foods properly can be a controlling factor.

Detection of Organism & Toxin Detection of Organism & Toxin

• Compendium of Methods for the • Toxin can be detected and typed by mouse Microbiological Examination of Foods. inoculation. • Enrichment of culture in cooked liver or cooked meat medium. • Plating on blood agar or egg yolk agar, Assumption: Active toxin will kill mouse and incubating anaerobically. • An ELISA method has been developed for detection of toxin.

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