Botulinum Antitoxin Research Articles

The Workshop on Animal Botulism in Europe

A workshop on animal botulism was held in Uppsala, Sweden, in June 2012. Its purpose was to explore the current status of the disease in Europe by gathering the European experts in animal botulism and to raise awareness of the disease among veterinarians and others involved in biopreparedness. Animal botulism is underreported and underdiagnosed, but an increasing number of reports, as well as the information gathered from this workshop, show that it is an emerging problem in Europe. The workshop was divided into 4 sessions: animal botulism in Europe, the bacteria behind the disease, detection and diagnostics, and European collaboration and surveillance. An electronic survey was conducted before the workshop to identify the 3 most needed discussion points, which were: prevention, preparedness and outbreak response; detection and diagnostics; and European collaboration and surveillance. The main conclusions drawn from these discussions were that there is an urgent need to replace the mouse bioassay for botulinum toxin detection with an in vitro test and that there is a need for a European network to function as a reference laboratory, which could also organize a European supply of botulinum antitoxin and vaccines. The foundation of such a network was discussed, and the proposals are presented here along with the outcome of discussions and a summary of the workshop itself.

Case report with sequential nerve conduction studies: A very old patient with atypical foodborne botulism type a with pupil sparing treated with botulinum antitoxin

Case report with sequential nerve conduction studies: A very old patient with atypical foodborne botulism type a with pupil sparing treated with botulinum antitoxin

Development of functional cell-based assays as replacement assays for botulinum toxins and antitoxins

Development of functional cell-based assays as replacement assays for botulinum toxins and antitoxins

Cranial neuropathies, confusion, and ataxia— Challenges for diagnosis and therapy

Introduction: The differential diagnosis of patients presenting with multiple cranial neuropathies, ataxia, and altered mentation is broad and includes immunologic, infectious, vasculitic and metabolic conditions. Primary considerations are Bickerstaff’s brainstem encephalitis (BBE), the Miller Fisher syndrome (MFS), Wernicke’s encephalopathy and botulism. The initial workup may be unrevealing. Timely treatment is imperative and unnecessary treatment can be associated with serious adverse reactions. Sensitivity to the decisions needed in such patients is therefore important. Case report: A 58-year-old male presented with symptoms of altered mental status, blurred vision, dysphagia and dysarthria, impaired pupillary responses to light, facial diplegia, ataxia, and decreased tendon reflexes after an episode of a self resolving diarrheal disease. Primary initial diagnostic concerns were Bickerstaff’s brainstem encephalitis (BBE), Miller Fisher syndrome (MFS), Wernicke’s encephalopathy and botulism. Initial work-up including cerebrospinal fluid analyses, imaging studies, and an electrodiagnostic examination did not provide information helpful for narrowing this differential. The patient was treated with botulinum antitoxin, thiamine and intravenous immune globulin (IVIG) before the results of specialized tests were available. The patient’s clinical condition improved. Retrospectively, the patient was diagnosed as BBE. Conclusion: This case emphasizes the difficulties in distinguishing between BBE, MFS, and botulism as well as demonstrating the complexities of treating such patients.

Intestinal Abscess Caused by Botulism

Introduction: Botulism is a rare but life threatening neuroparalytic condition caused by bacterium Clostridium botulinum. We report a case of a middle-aged male who developed intestinal abscesses that were secondary to Botulism. Case: A 43-year-old male was admitted to our institution with abdominal pain, blurred vision and dysarthria for one day duration. He denied any fever, headache or neck stiffness. Past medical history was significant for diabetes mellitus, hypertension and gout. Examination revealed normal vital signs, inability to protrude the tongue with no evidence of palpable adenopathy or rash. MRI brain was unremarkable. Laboratory values showed leukocytosis of 20.3 K/microliter and platelet count of 602 K/microliter. CT scan abdomen and pelvis showed multiloculated gas containing fluid collections in the perigastric area and in left anterior abdominal wall. Abscesses were incised and drained and culture from the abscess was positive for Clostridium botulinum type A toxin. Serum, stool and gastric aspirate were assayed using mouse bioassay which were positive for type A botulinum toxin. Subsequently, patient developed respiratory failure and was intubated requiring mechanical ventilatory support. The patient got botulinum antitoxin and was treated with imipenem. Subsequently, his condition started improving and is recovering neurologically in the last three weeks of follow-up. Discussion: Botulism is an acute neuroparalytic condition characterized by cranial nerve palsies followed by descending symmetrical flaccid paralysis which can cause respiratory arrest and can be fatal. Botulism is caused by botulinum neurotoxin types A, B, E or F and G produced by Clostridium species (C. botulism, C. baratti, C. butyricum). Three major forms of botulism are: 1) Food borne botulism from ingestion of food contaminated by preformed botulism neurotoxin. 2) Intestinal botulism caused by colonization from Clostridium botulinum followed by in situ production of toxin. 3) Wound botulism caused by contamination of wound by Clostridium botulinum and production of neurotoxin in situ. A careful history and physical examination, serum assays, analysis of stool and suspected food item may also reveal neurotoxin which is diagnostic when coupled with appropriate clinical and neurological findings. If clinical suspicion for botulism is high, Centers for Disease Control and Prevention (CDC) should be contacted for administration of botulism antitoxin which is the standard of care followed by antibiotics. To summarize, our case reiterates the importance of early and prompt diagnosis of botulism which is a fatal disease to initiate therapy in a timely fashion.

956 Infant Botulism Due to Honey Ingestation

Background and AimsBotulism is a neuroparalytic disease caused by neurotoxins produced by the bacteria Clostridium botulinum. This neurotoxin inhibits the normal release of acetylcholine in the synaptic cleft, inducing presynaptic...

Two cases of type A infant botulism in Grenoble, France: no honey for infants

We report two severe cases of infant botulism diagnosed at Grenoble University Hospital, France, respectively in 2006 and 2009. Both cases were characterized by a delay in diagnosis, severe neurological manifestations and extended period of hospitalization in intensive care unit, but a complete recovery. Infant botulism is a rare but life-threatening disease. It primarily affects infants, and the main risk factor is honey ingestion. Diagnosis should be systematically evoked by pediatricians in infants suffering from constipation, fatigue, muscle weakness, difficult feeding and altered cry, but before the onset of generalized flaccid paralysis, so as to administer specific treatment (BabyBIG®, a human derived botulinum antitoxin) at an early stage of the disease when it is most effective. In conclusion, parents should be aware of the role of honey as a source of spores of Clostridium botulinum and therefore infant botulism in the first year of life.

Fatal Course of Foodborne Botulism in an Eigth-Month Old Infant

An 8-month old girl, weighing 9 kg, was brought by her parents at 8.15 am to the Emergency Department (ED) for a progressive worsening of weakness and acute respiratory failure. On admission, the baby presented with poor oral intake, a weak cry and extremely weak muscular body control. Poor gag and suck, unreactive mydriasis, hypotonia, lethargy and absence of peristalsis were noted. Laboratory data showed severe respiratory acidosis. Chest X-ray, electroencephalography, encephalic CT scan and MRI were all normal, as were cerebrospinal fluid analysis and viral tests. Orotracheal intubation and continuous mechanical ventilation were applied. The patient received fluids, corticosteroids, aerosol therapy, large-spectrum antibiotics and enteral-nutrition. Further investigation revealed ingestion of an improperly prepared home-canned homogenized turkey meal. Type A botulinum neurotoxin was identified. Trivalent botulinum antitoxin, prostigmine and oral activated charcoal were administered. Generalized flaccid paralysis, areflexic bilateral mydriasis, gastric stasis and deep coma persisted for the duration of the hospital stay, and the patient died of severe respiratory failure and cardiac arrest 12 days after ED admission. Botulism poisoning should be suspected in any infant presenting with feeding difficulties, constipation, descendent paralysis or acute respiratory failure. Supportive treatment and antidotal therapy should be performed as soon as a clinical diagnosis is made. We describe a case of foodborne botulism in an 8-month old infant caused by ingestion of an improperly prepared home-canned homogenized turkey meal, representing the youngest fatal case reported in medical literature.

Initial Recovery and Rebound of Type F Intestinal Colonization Botulism After Administration of Investigational Heptavalent Botulinum Antitoxin

Investigational heptavalent botulinum antitoxin (HBAT) is now the primary antitoxin for US noninfant botulism patients. HBAT consists of equine Fab/F(ab')2 IgG fragments, which are cleared from circulation faster than whole immunoglobulins. Rebound botulism after antitoxin administration is not previously documented but occurred in our patient 10 days after HBAT administration.

Phrenic nerve-hemidiaphragm as a highly sensitive replacement assay for determination of functional botulinum toxin antibodies

Botulinum neurotoxins induce a prolonged muscle paralysis by specifically blocking the release of neuronal transmitters from peripheral nerve junctions. Potency testing of toxin and antitoxin therapies is entirely dependent on mouse lethality bioassay which is associated with extreme suffering of large numbers of animals to ensure high precision. The mouse phrenic nerve-diaphragm assay is an ex vivo assay that closely mimics in vivo respiratory paralysis offering substantial refinement and reduction in the number of animals used. A range of botulinum antitoxin standards, one licenced product and experimental antitoxins were tested for neutralising potency using ex vivo hemidiaphragm assay and compared with in vivo determined activities. Overall, there was an excellent agreement between neutralising activity detected by the two assay systems and for each toxin serotype using only 4–7 replicates for each product (almost perfect concordance for type A antitoxins: ρ = 0.997, and substantial concordance for type B antitoxins: ρ = 0.991 and type E antitoxins: ρ = 0.964, respectively). These findings confirm that the mouse nerve-diaphragm preparation can provide a functional ex vivo replacement assay for specific, sensitive and precise assessment of toxin and antitoxin activity.

Recurrent Wound Botulism Among Injection Drug Users in California

Botulism is an acute neurologic illness characterized by cranial nerve palsies and descending flaccid paralysis. Botulism is a rare disease and recurrent botulism even more rare. We review cases of recurrent wound botulism (WB) among injection drug users (IDUs) in California from 1993 through 2006 and describe 2 case patients. From botulism surveillance data for 1993-2006, we identified patients with >1 episode of clinical WB, defined as acute descending paralysis with a visible wound or recent history of injection drug use. For each patient, ≥1 of their WB episodes was laboratory confirmed. We extracted demographic, clinical, and laboratory information from case and laboratory reports and compared clinical characteristic frequency of initial and second WB episodes. During 1993-2006, 17 IDUs had recurrent WB, 14 with 1 recurrence and 3 with 2 recurrences. Of 25 laboratory-confirmed episodes, 22 were confirmed through serum testing and 3 through wound testing. Patients were 32-61 years old, and 94% were male. All patients reported heroin injections; 88% specified black tar heroin use and 76% reported subcutaneous injection. The most common presentations were having a visible wound, speech difficulty, double vision, respiratory difficulty, and trouble swallowing. There were no significant differences in clinical presentation between initial and second episodes. As the California epidemic of WB among IDUs continues, WB episodes are recurring. Both clinicians and IDUs should be aware of the potential for WB to recur among IDUs to enable timely diagnosis and early botulinum antitoxin administration and supportive care.

Neutralization of Tetanus Neurotoxin with Various Botulinum Antitoxins (A-F) in a Study to Isolate Clostridium botulinum from Field Samples

Neutralization of Tetanus Neurotoxin with Various Botulinum Antitoxins (A-F) in a Study to Isolate Clostridium botulinum from Field Samples

Infant botulism in Australia: availability of human botulinum antitoxin for treatment

We report the first Australian case of treatment of infant botulism with a human botulinum antitoxin developed in the United States by the California Department of Public Health. Our patient's clinical improvement was rapid, and although the product is expensive, cost-analysis supports the economical viability of its use. In future cases of suspected infant botulism, we recommend that Australian clinicians promptly obtain and administer this antitoxin to their patient.

Rare Case of Dysphagia

Wound botulism is exceedingly rare and occurs almost exclusively among injection drug users. In 2008 there was a case of wound botulism in a noninjecting drug user reported to the Centers for Disease Control and Prevention (CDC). We report a case of a Caucasian male developing dysphagia due to wound botulism after having a motorcycle accident that left him with open fractures. The CDC was contacted and the patient was transferred to medical intensive care unit to be emergently started on hepatavalent Clostridium botulinum antitoxin. Early suspicion of wound botulism is essential for effective therapy with antitoxin in this life-threatening disease. If not suspected, this patient would likely have died. Nevertheless, the delay in diagnosis and treatment resulted in the patient's suffering dysphagia and neurological deficits. The patient required a percutaneous endoscopic gastrostomy tube and months of dysphagia therapy, supportive care, and rehabilitation. Our aim is to increase the awareness for wound botulism when a patient presents with dysphagia and diplopia after suffering open wounds. If suspected early, the morbidity and mortality from this disease can be prevented.

Source of Botulinum Antitoxin Products Needs Clarifying

Source of Botulinum Antitoxin Products Needs Clarifying

Oropharyngeal Dysphagia in Wound Botulism

AbstractBotulism results in a symmetric bulbar paralysis due to neuroparalytic toxins produced by Clostridium botulinum. These toxins block release of the neurotransmitter acetylcholine at the neuromuscular junction, resulting in paralysis. Several forms of botulism occur, all rare in the United States. The prominent bulbar involvement suggests oropharyngeal dysphagia is common in botulism. This article reviews the botulism literature with an emphasis on dysphagia, the swallowing signs, and clinical course in a single case of noninjection drug use wound botulism. The patient was admitted to an outside hospital with dysphagia and dysarthria 11 days after suffering a traumatic sports injury. He was later transferred to our tertiary care facility, diagnosed with wound botulism, and administered botulinum antitoxins. Repeated clinical swallow examinations were notable for decreased hyolaryngeal excursion, coughing after swallowing, and poor secretion management. Eight days after administration of botulinum an...

Persistence of Botulinum Toxin in Patients' Serum: Alaska, 1959–2007

Persistence of circulating toxin in patients with foodborne botulism is not well characterized. Recommendations for administration of botulinum antitoxin are ambiguous for patients with late-presenting disease, such as a Florida woman with toxin-positive serum 12 days after toxin ingestion. We reviewed Alaska records of foodborne outbreaks of botulism that occurred during 1959-2007 to examine the period after ingestion during which toxin was detected. Of 64 cases with toxin-positive serum, toxin was detected up to 11 days after ingestion. The findings from Alaska and Florida support administration of antitoxin up to 12 days after toxin ingestion but do not indicate when circulating toxin should no longer be present.

Measurement of botulinum types A, B and E neurotoxicity using the phrenic nerve–hemidiaphragm: Improved precision with in-bred mice

Botulinum neurotoxins induce a prolonged muscle paralysis by specifically blocking the release of neuronal transmitters from peripheral nerve junctions. The current method for assessing the potency of botulinum toxin and antitoxins is the mouse LD 50 assay. The mouse phrenic nerve–diaphragm assay is an in vitro assay that closely mimics in vivo respiratory paralysis. In this study, we have further improved the assay by using gelatin as a non-frothing alternative to albumin and investigated the effects of botulinum toxin serotypes A, B and E on phrenic nerve–hemidiaphragms from out-bred MF1 and in-bred Balb/c mice. Improved reproducibility was found with in-bred mice. Balb/c mice were also found to be much less sensitive to type B toxin perhaps indicating differences in the expression of receptor components. Hemidiaphragm preparations from Balb/c mice were approximately 7 times more sensitive to type A toxin and 7–12 times more sensitive to type E toxin relative to type B toxin. These findings indicate that when fully optimised the mouse nerve–diaphragm preparation can provide a functional in vitro model for accurate and reproducible assessment of toxin activity.

Wound botulism from heroin skin popping

Following the introduction of black tar heroin mainly from Mexico in the 1980s, cases of wound botulism dramatically increased in the western United States. Contamination with spores of Clostridium botulinum of black tar heroin occurs along the distribution line. The heating of heroin powder to solubilize it for subcutaneous injection ("skin popping") does not kill the spores. The spores germinate in an anaerobic tissue environment and release botulinum toxin type A or B. Unless skin abscesses are found in the patient, the clinical diagnosis is often challenging. Facilitation of the compound muscle action potential by repetitive nerve stimulation at 20 to 50 Hz is an important and rapid diagnostic test. Definite diagnosis is made by detection of botulinum toxin in serum or isolation of C botulinum from the abscess. Early treatment with equine ABE botulinum antitoxin obtained from the Centers for Disease Control and Prevention often shortens the time on a ventilator.

Process development and pilot-scale production of therapeutic antisera against botulinum toxin type A and B

This study aimed at process development and pilot-scale production of botulinum antitoxin for an urgent botulism-poisoning patient, contributing to steady antitoxin supply for bio-terrorism. After horses were immunized with formalin-inactivated botulinum toxin type A and B, the sera were prepared. Following purifications of IgG fractions by cold ethanol fractionation, caprylate fractionation, anion and exchange chromatography, and F(ab′)2 by cold ethanol fractionation, pepsin digestion, and cation exchange chromatography, the specificity and protective efficacy against botulinum toxin type A and B were analyzed by enzyme-linked immunosorbent assay (ELISA), Western blot, and mouse neutralization assay. Based on the result of mouse neutralization assay, it was found that 25 μg of anti-type A IgG and 350 μg of anti-type B IgG were equal to 1 IU. Neutralizing potency of F(ab′)2 was also analyzed as 30 and 350 μg/IU for type A and B toxins, respectively, showing that the neutralizing activity of the F(ab′)2 is close to that of IgGs. In this report, we successfully established the manufacturing process of equine botulinum toxin type A and B antitoxin and produced freeze-dried prototype products.