Loretta M. McCroskey

An Outbreak of Type A Botulism Associated with a Commercial Cheese Sauce

Botulism is a rare disease; between 1983 and 1992, an average of only 22 cases of food-borne botulism were reported to the Centers for Disease Control and Prevention each year (Unpublished data). Nevertheless, a reported case of food-borne botulism represents a public health emergency because many persons may be affected if the contaminated food is not identified. This is especially true of outbreaks linked to commercial products or restaurants. However, because few clinicians have ever seen a case of botulism, the diagnosis may be delayed or not even considered. Diagnosing botulism is a special challenge when patients present with mild symptoms and do not have a history of exposure to typical food vehicles, such as home-canned vegetables. We describe an outbreak of botulism that was characterized by relatively mild symptoms and subtle physical findings. The outbreak was caused by a food vehicle that was initially considered to be unlikely. The investigation shows the importance of considering the diagnosis of botulism soon after patients present with acute cranial nerve dysfunction and of promptly reporting suspected cases to public health officials. The Outbreak On 4 October 1993, a 42-year-old woman (patient 1) visited her family physician in a small town in southern Georgia. She had had nausea, blurred vision, and loss of balance for 2 days. Results of physical examination were normal except for a possible sixth-nerve palsy. Labyrinthitis was diagnosed, and the patient was sent home. When her physician contacted her the next day, she was too weak to come to the telephone. Her husband reported that her speech was slurred and that she was having difficulty swallowing. He mentioned that their 21-year-old daughter (patient 2) also had nausea and difficulty swallowing. Both patients were referred to a neurologist, who recognized this unusual clustering of neurologic symptoms as possible botulism. The patients were admitted to a hospital, and public health officials were notified. That same day, a 38-year-old woman with a history of hypertension (patient 3) was seen in the emergency department of the same hospital because of blurred vision, slurred speech, and weakness in her right arm. She was admitted to the medical ward with a diagnosis of transient ischemic attack. By coincidence, the family physician of patients 1 and 2 was also attending on the medical ward that night; he recognized that patient 3 might be another case of botulism. Patient 3 mentioned that her friend, patient 4, was having similar symptoms. Patient 4 was notified that her illness might also be botulism, and she too was hospitalized. Two days earlier, her new symptoms had been diagnosed as an allergic reaction to a tranquilizer. Patient 5 had visited an optometrist on 5 October with fatigue and blurred, double vision. She received a diagnosis of mild glaucoma and astigmatism and was given a prescription for eyeglasses. She presented to the emergency department on 7 October after hearing about the outbreak on the radio. None of the patients had eaten any home-canned foods. However, on 1 October, all of them had eaten food from a delicatessen that had re-opened on 23 September after having been closed for 6 months because of the owners family obligations. Local health officials closed the delicatessen on 6 October and seized leftover foods. Methods Clinical and Epidemiologic Investigation Hypothesis-generating interviews were done with the hospitalized patients and the owner of the delicatessen. Each step in the preparation and storage of foods was reviewed. After a standardized questionnaire that addressed food histories and symptoms was developed, investigators attempted to interview (either by telephone or in person) all persons who had eaten food from the delicatessen in the 6 days it was open between 23 September and 2 October 1993. For the purposes of the investigation, a case of botulism was defined as dysphagia, dysphonia, dysarthria, or diplopia that developed after 23 September in any person who had eaten food purchased at the delicatessen. A press release was issued to identify patrons of the delicatessen. The press release asked all persons who had eaten at the delicatessen to call the local health department. The owner and known patrons of the delicatessen were asked to name other patrons, and businesses in the neighborhood around the delicatessen were surveyed as to whether workers had eaten food from the delicatessen. In an attempt to find additional cases, 50 physicians in the area were called and asked whether they had seen any patients since 23 September who reported blurred or double vision, dry mouth, difficulty swallowing, change in voice, or muscle weakness. To identify any cases that may have been mistakenly diagnosed as other conditions, physicians were also asked if they had recently seen any patients with a diagnosis of stroke, transient ischemic attack, the Guillain-Barre syndrome, or myasthenia gravis. Logs from the emergency department of the local hospital were reviewed for these symptoms and diagnoses. All hospitalized patients were examined by the same neurologist, and their hospital and outpatient records were reviewed. No neurologic examination was done on three persons who met the case definition for botulism but did not seek medical attention. These patients were identified by their responses to the standard questionnaire. The delicatessen was inspected by officials of the Georgia Department of Agriculture. Officials of the Food and Drug Administration inspected the canning facility and searched for unused cans of the same batch of cheese sauce. Laboratory Investigation Samples of food taken from the delicatessen were assayed for botulinum toxin and were cultured for Clostridium botulinum as described elsewhere [1]. All persons who ate the implicated food were asked to submit serum and stool specimens. Gastric aspirate specimens were obtained from two hospitalized patients. Serum, stool, and gastric aspirate specimens were assayed for C. botulinum toxin, and stool specimens were cultured for C. botulinum. Inoculation experiments were done in the Food and Drug Administration botulism laboratory to determine the time and temperature needed for C. botulinum to grow and for toxin to be produced in the implicated brand of cheese sauce. Spores harvested from cultures of the outbreak strain of C. botulinum were heat-shocked at 80 C for 10 minutes and then diluted with sterile water to a concentration of 104 spores/mL. Twenty g of the cheese sauce was then added to sterile test tubes that contained 0.1 mL of inoculum; the final concentration was 103 spores/20 g of cheese. The tubes were incubated at 22 C and 5 C; they were then assayed for toxin on day 8 and every 3 to 4 days for 2 months. Toxin testing was also done before incubation to ensure that no toxin was transferred with the inoculum. Toxin was measured in mouse minimum lethal doses using the mouse bioassay [2]. Results Epidemiologic Findings The delicatessen first opened in August 1992 and then closed for 6 months from March to September 1993. It reopened on 23 September 1993, serving lunch 3 days a week (Thursday through Saturday). Food was served at the delicatessen from 23 to 25 September and from 30 September to 2 October (Figure 1). Routine inspections of the delicatessen done on 30 September and after the outbreak showed no violations of state standards for retail food sale establishments. Figure 1. Date of meals eaten by all 52 patrons of the delicatessen and dates of symptom onset in 8 patrons with botulism. Fifty-two persons who ate food from the delicatessen in the 6 days it was open between 23 September and 2 October 1993 were identified and interviewed. Eight (15%) met the case definition for botulism. Their ages ranged from 20 to 48 years; 6 were women. No additional cases were identified through the review of emergency department logs or the physician survey. The owner of the delicatessen estimated that she served about 20 meals each day. Many of the patrons were friends of the owner or members of the owners family and had eaten there more than once. Eight (36%) of 22 persons who ate food from the delicatessen on Friday, 1 October, met the case definition compared with none of the 30 who ate the food only on other days. Among the 22 persons who ate food from the delicatessen on 1 October, all 8 ill persons but none of 14 well persons had eaten a barbecue stuffed potato. Six other persons had eaten barbecue stuffed potatoes before 1 October but remained well. The owner did not recall selling any stuffed potatoes on 2 October. Clinical Findings and Laboratory Confirmation of Botulism The illnesses ranged from mild to severe. Most patients had few, subtle objective neurologic findings despite having many symptoms characteristic of botulism. Symptoms developed a median of 2.5 days after exposure (range, 1 to 6 days). All ill persons had 3 or more symptoms consistent with botulism (median, 8.5 symptoms; range, 3 to 14 symptoms) and, by definition, at least 1 symptom that suggested a cranial nerve abnormality. The most common symptoms were dry mouth, difficulty speaking and swallowing, and change in voice quality (Table 1). All ill persons had neurologic and gastrointestinal symptoms. Three persons had illnesses so mild that they did not seek medical care. Five persons were hospitalized in an intensive care unit after botulism was suspected; in four of these persons, an illness other than botulism was initially diagnosed (Table 2). Patient 3, the most severely affected, developed complete bilateral ptosis, markedly dysarthric speech, weakness of the tongue and palate, arm and leg weakness, and respiratory failure. She died of a pulmonary embolism after being supported by mechanical ventilation for 18 days. Patient 1 was noticeably dysarthric and required nasogastric intubation because of difficulty swallowing. Patients 2, 4, and 5 had few object

Genetic Homogeneity of Clostridium botulinum Type A1 Strains with Unique Toxin Gene Clusters

ABSTRACT A group of five clonally related Clostridium botulinum type A strains isolated from different sources over a period of nearly 40 years harbored several conserved genetic properties. These strains contained a variant bont/A1 with five nucleotide polymorphisms compared to the gene in C. botulinum strain ATCC 3502. The strains also had a common toxin gene cluster composition (ha−/orfX+) similar to that associated with bont/A in type A strains containing an unexpressed bont/B [termed A(B) strains]. However, bont/B was not identified in the strains examined. Comparative genomic hybridization demonstrated identical genomic content among the strains relative to C. botulinum strain ATCC 3502. In addition, microarray data demonstrated the absence of several genes flanking the toxin gene cluster among the ha−/orfX+ A1 strains, suggesting the presence of genomic rearrangements with respect to this region compared to the C. botulinum ATCC 3502 strain. All five strains were shown to have identical flaA variable region nucleotide sequences. The pulsed-field gel electrophoresis patterns of the strains were indistinguishable when digested with SmaI, and a shift in the size of at least one band was observed in a single strain when digested with XhoI. These results demonstrate surprising genomic homogeneity among a cluster of unique C. botulinum type A strains of diverse origin.

Detection and differentiation of Clostridium botulinum type A strains using a focused DNA microarray

A focused oligonucleotide microarray featuring 62 probes targeting strain variable regions of the Clostridium botulinum strain ATCC 3502 genome sequence was developed to differentiate C. botulinum type A strains. The strain variable regions were selected from deletions identified among a panel of 10 type A strains compared to the strain ATCC 3502 genome sequence using high density comparative genomic hybridization microarrays. The focused microarray also featured specific probes for the detection of the neurotoxin genes of various serotypes (A-G), toxin gene cluster components (ha70 and orfX1), and fldB as a marker for proteolytic clostridia (Group I). Eight pairs of strains selected from separate type A botulism outbreaks were included in the 27 subtype A1-A4 strains examined in this study. Each outbreak related strain pair consisted of strains isolated from different sources (stool and food). Of the eight outbreak related strain pairs, six groups of strains with indistinguishable hybridization patterns were identified. Outbreak related strains were shown to have identical hybridization patterns. Strain pairs from three separate outbreaks involving strains harboring both the type A neurotoxin gene (bont/A) and an unexpressed type B neurotoxin gene (bont/B) shared the same probe hybridization profile. The focused microarray format provides a rapid approach for neurotoxin gene detection and preliminary determination of the relatedness of strains isolated from different sources.

Laboratory Investigation of the First Case of Botulism Caused by Clostridium butyricum Type E Toxin in the United States

ABSTRACT We report here the laboratory investigation of the first known case of botulism in the United States caused by Clostridium butyricum type E. This investigation demonstrates the importance of extensive microbiological examination of specimens, which resulted in the isolation of this organism.