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Botulinum toxin is a protein toxin secreted by Clostridium botulinum that is strongly neurotoxic. Due to its characteristics of being super toxic, quick acting, and difficult to prevent, the currently reported antiviral studies focusing on monoclonal antibodies have limited effectiveness. Therefore, for the sake of effectively prevention and treatment of botulism and to maintain country biosecurity as well as the health of the population, in this study, we intend to establish a single chain antibody (scFv) targeting the carboxyl terminal binding functional domain of the botulinum neurotoxin heavy chain (BONT/AHc) of botulinum neurotoxin type A, and explore the value of a new passive immune method in antiviral research which based on adeno-associated virus (AAV) mediated vector immunoprophylaxis (VIP) strategy. The scFv small-molecular single-chain antibody sequenced, designed, constructed, expressed and purified by hybridoma has high neutralising activity and affinity level, which can lay a good foundation for the modification and development of antibody engineering drugs. In vivo experiments, AAV-mediated scFv engineering drug has good anti-BONT/A toxin neutralisation ability, has advantages of simple operation, stable expression and good efficacy, and may be one of the effective treatment strategies for long-term prevention and protection of BONT/A botulinum neurotoxin.

Botulism is one of the most serious food intoxications, manifesting as prolonged paralytic conditions. This disease is usually the result of the consumption of poor quality canned or smoked foods, so the inhabitants of many countries of the world are exposed to the risk of this kind of poisoning every year. In view of the severity of poisonings caused by botulinum neurotoxins, monoclonal antibodies (mAbs) show great promise because of their targeting action, lack of allergic reactions and serum sickness. The use of a cocktail of mAbs increases the "functional specificity" of their mixture, allowing them to bind to the active domains of different toxin chains and block their action. In this work, we obtained 14 murine mAbs to the catalytic and receptor-binding domain of botulinum toxin type A. The Sp2/0-Ag14 murine myeloma cell line and splenocytes from immunized mice of the BALB/c line were used as fusion partners. We have shown that the selected cocktail of three antibodies neutralizes native toxin more effectively than antibodies separately-complete neutralization is achieved at a toxin dose of 3LD50 and partial neutralization at 5LD50. We presume that this cocktail may be promising as a prototype for the creation of a therapeutic drug capable of neutralizing the toxin in the blood of patients.

Introduction: Iatrogenic botulism is a rare, serious disease that progresses with descending paralysis and develops after cosmetic or therapeutic botulinum toxin-A (BoNT-A) application., Case Presentations: In this case series; six cases of iatrogenic botulism followed up in our center are presented. Four of these developed after gastric BoNT-A and two after axillary BoNT-A application., Results: The most important cause for the disease was the use of unlicensed products and high-dose toxin applications. The first symptoms were blurred vision, double vision, difficulty in swallowing, and hoarseness. Symptoms appeared within 4-10 days after the application of BoNT-A. Symptoms progressed in the course of descending paralysis in the following days with fatigue, weakness in extremities and respiratory distress. Diagnosis was based on patient history and clinical findings. The main principles of foodborne botulism therapy were applied in the treatment of iatrogenic botulism. If clinical worsening continued, regardless of the time elapsed after BoNT-A application, the use of botulinum antitoxin made a significant contribution to clinical improvement and was recommended., Conclusions: Routine and new indications for BoNT-A usage are increasing and, as a result, cases of iatrogenic botulism will be encountered more frequently. Physicians should be alert for iatrogenic botulism in the follow-up after BoNT-A applications and in the differential diagnosis of neurological diseases that are presented with similar findings., Competing Interests: No Conflict of Interest is declared, (Copyright (c) 2024 Fatma Eser, İmran Hasanoğlu, Bircan Kayaaslan, Ayşe Kaya Kalem, Şule Bilen, Gürdal Orhan, Rahmet Güner.)

Equine botulinum antitoxin is one of the most popular countermeasures for human botulism. The unitage of the antitoxin product is defined according to national minimum requirement or pharmacopoeia in each country by referring to national standard antitoxins for four types (A, B, E, and F). With the expected depletion of the national standard antitoxins, replacement national standard antitoxins are produced and standardized through collaboration of the National Control Laboratory and other participants, including manufacturer(s). Therefore, Japanese National Standard Botulinum Antitoxin Type A, Equine, was replaced according to the results of a collaborative study involving the National Institute of Infectious Diseases and KM Biologics Co., Ltd. The unitage of the replacement material was determined through mouse neutralization tests, which involved toxin-antitoxin mixture injection at pH 7.0. Potency value of 440 units/vial was obtained. However, the Japanese Minimum Requirement for Biological Products was revised, and the neutralization reactions were repeated at pH 6.0, for which considerably different potency value (656 units/vial) and survival profile of mice were obtained. In September 2021, the replacement material, Japanese National Standard Botulinum Antitoxin Type A, Equine, lot 2, was established with potency value of 656 Units/vial. The impact of pH-dependent change in potency on antitoxin quality control is discussed.

Botulinum neurotoxin (BoNT) is a potent protein toxin that causes muscle paralysis and death by asphyxiation. Treatments for symptomatic botulism are intubation and supportive care until respiratory function recovers. Aminopyridines have recently emerged as potential treatments for botulism. The clinically approved drug 3,4-diaminopyridine (3,4-DAP) rapidly reverses toxic signs of botulism and has antidotal effects when continuously administered in rodent models of lethal botulism. Although the therapeutic effects of 3,4-DAP likely result from the reversal of diaphragm paralysis, the corresponding effects on respiratory physiology are not understood. Here, we combined unrestrained whole-body plethysmography (UWBP) with arterial blood gas measurements to study the effects of 3,4-DAP, and other aminopyridines, on ventilation and respiration at terminal stages of botulism in mice. Treatment with clinically relevant doses of 3,4-DAP restored ventilation in a dose-dependent manner, producing significant improvements in ventilatory parameters within 10 minutes. Concomitant with improved ventilation, 3,4-DAP treatment reversed botulism-induced respiratory acidosis, restoring blood levels of CO 2 , pH, and lactate to normal physiologic levels. Having established that 3,4-DAP-mediated improvements in ventilation were directly correlated with improved respiration, we used UWBP to quantitatively evaluate nine additional aminopyridines in BoNT/A-intoxicated mice. Multiple aminopyridines were identified with comparable or enhanced therapeutic efficacies compared with 3,4-DAP, including aminopyridines that selectively improved tidal volume versus respiratory rate and vice versa. In addition to contributing to a growing body of evidence supporting the use of aminopyridines to treat clinical botulism, these data lay the groundwork for the development of aminopyridine derivatives with improved pharmacological properties. SIGNIFICANCE STATEMENT: There is a critical need for fast-acting treatments to reverse respiratory paralysis in patients with botulism. This study used unrestrained, whole-body plethysmography and arterial blood gas analysis to show that aminopyridines rapidly restore ventilation and respiration and reverse respiratory acidosis when administered to mice at terminal stages of botulism. In addition to supporting the use of aminopyridines as first-line treatments for botulism symptoms, these data are expected to contribute to the development of new aminopyridine derivatives with improved pharmacological properties., (Copyright © 2024 by The Author(s).)

Background: Rosacea is a chronic inflammatory disease usually associated with persistent erythema and periodic flushing. This disease is difficult to treat, and the outcomes are often unsatisfactory and prone to recurrence. In recent years, botulinum toxin has been used as a new treatment for rosacea; however, its efficacy and safety remain under discussion. Although a systematic review of the effectiveness and safety of botulinum toxin has been previously conducted by other researchers, our systematic review and meta-analysis evaluate the efficacy of botulinum toxin from a more comprehensive and detailed perspective to provide evidence for clinicians., Methods: Any study using botulinum toxin for the treatment of rosacea was considered for the analysis., Results: A total of 22 studies were included, 9 of which were randomized controlled trials involving 720 subjects. After treatment, all studies showed varying degrees of improvement in patient signs and symptoms along with reduced Clinician's Erythema Assessment (CEA) scores. The improvement was maintained for several months, and the adverse effects were mild and self-limiting., Conclusion: Botulinum toxin may be an effective treatment for patients with rosacea; however, further clinical evidence is needed to confirm its long-term efficacy and side effects. The study was preregistered with Prospero (CRD42022358911)., (© 2023 The Authors. Journal of Cosmetic Dermatology published by Wiley Periodicals LLC.)

Clostridium botulinum toxin-A (BoNT-A) creates temporary paralysis in the muscles by acting on the muscle-nerve junction. It is injected into the mimic muscles when a decrease in the movements of the mimic muscles is desired. Despite many favorable applications, the use of BoNT-A is not without drawbacks. Although there is no expected serious side effect on health in BoNT-A treatments, various problems can be encountered in patients treated for aesthetic purposes. Botulism is a rare but potentially life-threatening syndrome, which is caused by the toxin produced by the bacterium Clostridium botulinum , which acts on the nervous system, vegetative forms of C. botulinum can only survive in anaerobic conditions, while spore forms are common in nature and can withstand harsh conditions. Botulism can stem from bacterial spores which release toxin in the body; in the form of enteric botulism, and wound botulism. The cases that develop 'iatrogenic botulism' after such procedures are usually those receiving high-dose toxin for therapeutic purposes. The treatment of botulism mainly consists of anti-toxin therapy and, if necessary, intensive care to prevent organ failures, including respiratory support. This article aims to cover all these issues related to botulism and other adverse outcomes related to BoNT-A injection in light of the most recent literature., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Clostridium botulinum neurotoxin type A (BoNT/A) is one of the most potent biotoxins ever known. Its entry into neurons could block vesicle exocytosis to abolish the release of neurotransmitters from nerve terminals, thus leading to muscle paralysis. Although there are so many peptides, antibodies and chemical compounds claimed to have anti-toxin activity, no drug is available in the clinical application except equine antitoxin serum. In the present work, a short peptide inhibitor RRGW of BoNT/A was firstly identified by computer-aided ligand-receptor binding simulation, then an RRGW derived peptide was rational designed based on the fragment of SNAP-25 (141-206 aa). Proteolytic assay showed that the anti-toxin activity of the RRGW derived peptide was much higher than that of RRGW. Digit abduction score assay demonstrated that the derived peptide delayed BoNT/A-induced muscle paralysis at a lower concentration by 20-fold than RRGW. The results supported that RRGW derived peptide can be a potential BoNT/A inhibitor candidate for further treating botulism.

Objective: Botulinum toxin type A is widely used for the treatment of spasticity, focal dystonia, hemifacial spasm, hyperhidrosis, strabismus, chronic migraine, and also cosmetic purposes. Therapeutic use is commonly effective and safe. However, if toxin enters the vascular space and gets through to peripheral cholinergic nerve terminals, it may lead to iatrogenic botulism., Method: We presented a patient who is diagnosed as iatrogenic botulism and treated with antitoxin at the 15th day of the exposure., Results: After the antitoxin administration, dramatical response to the treatment was observed., Conclusions: In this report, we want to evaluate a new case of iatrogenic botulism and emphasize the importance of antitoxin administration regardless the timing of the exposure for patients with progressing paralysis., Competing Interests: Conflicts of Interest and Source of Funding: The authors have no conflicts of interest to declare., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Botulism is a low incidence but potentially fatal infectious disease caused by neurotoxins produced mainly by Clostridium botulinum . There are different routes of acquisition, food-borne and infant/intestinal being the most frequent presentation, and antitoxin is the treatment of choice in all cases. In Spain, botulism is under surveillance, and case reporting is mandatory., Methods: This retrospective study attempts to provide a more complete picture of the epidemiology of botulism in Spain from 1997 to 2019 and an assessment of the treatment, including the relationship between a delay in antitoxin administration and the length of hospitalization using the Cox proportional hazards test and Kruskal-Wallis test, and an approach to the frequency of adverse events, issues for which no previous national data have been published., Results: Eight of the 44 outbreaks were associated with contaminated commercial foods involving ≤7 cases/outbreak; preserved vegetables were the main source of infection, followed by fish products; early antitoxin administration significantly reduces the hospital stay, and adverse reactions to the antitoxin affect around 3% of treated cases.

Botulinum neurotoxin serotype A (BoNT/A) is recognized by the Centers for Disease Control and Prevention (CDC) as the most potent toxin and as a Tier 1 biowarfare agent. The severity and longevity of botulism stemming from BoNT/A is of significant therapeutic concern, and early administration of antitoxin-antibody therapy is the only approved pharmaceutical treatment for botulism. Small molecule therapeutic strategies have targeted both the heavy chain (HC) and the light chain (LC) catalytic active site and α-/β-exosites. The LC translocation mechanism has also been studied, but an effective, nontoxic inhibitor remains underexplored. In this work, we screened a library of salicylanilides as potential translocation inhibitors. Potential leads following a primary screen were further scrutinized to identify sal 30 , which has a cellular minimal concentration of a drug that is required for 50% inhibition (IC 50 ) value of 141 nM. The inquiry of salicylanilide sal 30 's mechanism of action was explored through a self-quenched fluorogenic substrate conjugated to bovine serum albumin (DQ-BSA) fluorescence, confocal microscopy, and vacuolar H + -ATPase (V-ATPase) inhibition assays. The summation of these findings imply that endolysosomal proton translocation through the protonophore mechanism of sal 30 causes endosome pH to increase, which in turn prevents LC translocation into cytosol, a process that requires an acidic pH. Thus, the inhibition of BoNT/A activity by salicylanilides likely occurs through disruption of pH-dependent endosomal LC translocation. We further probed BoNT inhibition by sal 30 using additivity analysis studies with bafilomycin A1, a known BoNT/A LC translocation inhibitor, which indicated the absence of synergy between the two ionophores.

Botulism antitoxin heptavalent (A, B, C, D, E, F, and G - Equine; BAT) product is a sterile solution of F(ab') 2 and F(ab') 2 -related antibody fragments prepared from plasma obtained from horses that have been immunized with a specific serotype of botulinum toxoid and toxin. BAT product is indicated for the treatment of symptomatic botulism following documented or suspected exposure to botulinum neurotoxin serotypes A to G in adults and pediatric patients. Pharmacokinetic and exposure-response models were used to explore the relationship between BAT product exposure and the probability of survival, and the occurrence of relevant moderate clinical signs observed during the preclinical development of BAT product to justify the clinical dose. The predicted probability of survival in humans for all serotypes of botulinum neurotoxin was more than 95.9% following intravenous administration of one vial of BAT product. Furthermore, this BAT product dose is expected to result in significant protection against clinical signs in human adults for all botulinum neurotoxin serotypes. Our exposure response model indicates that we have sufficient antitoxin levels to give full protection at various theoretical exposure levels and, based on neutralization capacity/potency of one dose of BAT product, it is expected to exceed the amount of circulating botulinum neurotoxin., (© 2022 Emergent Biodefense. Clinical Pharmacology & Therapeutics published by Wiley Periodicals LLC on behalf of American Society for Clinical Pharmacology and Therapeutics.)

Botulinum neurotoxins (BoNTs) are highly potent, select agent toxins that inhibit neurotransmitter release at motor nerve terminals, causing muscle paralysis and death by asphyxiation. Other than post-exposure prophylaxis with antitoxin, the only treatment option for symptomatic botulism is intubation and supportive care until recovery, which can require weeks or longer. In previous studies, we reported the FDA-approved drug 3,4-diaminopyridine (3,4-DAP) reverses early botulism symptoms and prolongs survival in lethally intoxicated mice. However, the symptomatic benefits of 3,4-DAP are limited by its rapid clearance. Here we investigated whether 3,4-DAP could sustain symptomatic benefits throughout the full course of respiratory paralysis in lethally intoxicated rats. First, we confirmed serial injections of 3,4-DAP stabilized toxic signs and prolonged survival in rats challenged with 2.5 LD 50 BoNT/A. Rebound of toxic signs and death occurred within hours after the final 3,4-DAP treatment, consistent with the short half-life of 3,4-DAP in rats. Based on these data, we next investigated whether the therapeutic benefits of 3,4-DAP could be sustained throughout the course of botulism by continuous infusion. To ensure administration of 3,4-DAP at clinically relevant doses, three infusion dose rates (0.5, 1.0 and 1.5 mg/kg∙h) were identified that produced steady-state serum levels of 3,4-DAP consistent with clinical dosing. We then compared dose-dependent effects of 3,4-DAP on toxic signs and survival in rats intoxicated with 2.5 LD 50 BoNT/A. In contrast to saline vehicle, which resulted in 100% mortality, infusion of 3,4-DAP at ≥ 1.0 mg/kg∙h from 1 to 14 d after intoxication produced 94.4% survival and full resolution of toxic signs, without rebound of toxic signs after infusion was stopped. In contrast, withdrawal of 3,4-DAP infusion at 5 d resulted in re-emergence of toxic sign and death within 12 h, confirming antidotal outcomes require sustained 3,4-DAP treatment for longer than 5 d after intoxication. We exploited this novel survival model of lethal botulism to explore neurophysiological parameters of diaphragm paralysis and recovery. While neurotransmission was nearly eliminated at 5 d, neurotransmission was significantly improved at 21 d in 3,4-DAP-infused survivors, although still depressed compared to naïve rats. 3,4-DAP is the first small molecule to reverse systemic paralysis and promote survival in animal models of botulism, thereby meeting a critical treatment need that is not addressed by post-exposure prophylaxis with conventional antitoxin. These data contribute to a growing body of evidence supporting the use of 3,4-DAP to treat clinical botulism., (© 2022. The Author(s).)

The review summarises the current knowledge of the treatment of iatrogenic botulinum toxin overdose. The symptoms may be diffuse, and suspicion should be raised based on time of symptom appearance relative to the time of exposure. Iatrogenic botulism may appear if the maximum recommended total dose of botulinum toxin has been exceeded and if the drug is spread locally from the site of injection or is redistributed to the systemic circulation. The adverse drug reactions frequency is possibly underreported. Fast initiation of the available antidote may be needed. The guideline provided on treatment of iatrogenic botulism is developed from non-iatrogenic botulism.

Competing Interests: Financial disclosure statements have been obtained, and no conflicts of interest have been reported by the authors or by any individuals in control of the content of this article.

In October 2019, ninety-four patients were admitted into Alexandria Poison Center (APC) with a history of ingestion of Feseekh (salted fish). In an attempt to allocate the resources, not all patients were given HBAT (botulism antitoxin heptavalent (A, B, C, D, E, F, G) equine immediately. The current study aimed to portray the clinical characteristics of the cases, explore the possible relation between these characteristics and necessity of HBAT administration, explore the reliability of mouse lethal test, and establish a clinical guide for management including preservation of resources. The current prospective study included 94 patients who were admitted to Alexandria Poison Center (APC) in the period from the 29 th of September to the 27 th of October 2019. The patients' data were recorded using a checklist that includes: personal data, past medical history, clinical assessment, investigations, treatment, and the outcome. The checklist was carried out to assess and follow up each patient. Hospitalized patients were categorized according to symptoms consistent with botulism. The equine HBAT, made by Emergent BioSolutions Canada Inc. (formerly Cangene Corporation), was used in the treatment. HBAT was given to thirty-four patients (36.2%) only out of the total admission. However, eighty-two (87.2%) of patients were completely cured, whereas ten patients (10.6%) were discharged with mild neurological sequels and death occurred only in two cases (2.2%). Sixty cases (63.8%) with suspected foodborne botulism could be managed by supportive treatment only with no need for HBAT, while patients with evident neurological signs received HBAT immediately., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Antitoxin, the only licensed drug therapy for botulism, neutralizes circulating botulinum neurotoxin (BoNT). However, antitoxin is no longer effective when a critical amount of BoNT has already entered its target nerve cells. The outcome is a chronic phase of botulism that is characterized by prolonged paralysis. In this stage, blocking toxin activity within cells by next-generation intraneuronal anti-botulinum drugs (INABDs) may shorten the chronic phase of the disease and accelerate recovery. However, there is a lack of adequate animal models that simulate the chronic phase of botulism for evaluating the efficacy of INABDs. Herein, we report the development of a rabbit model for the chronic phase of botulism, induced by intoxication with a sublethal dose of BoNT. Spirometry monitoring enabled us to detect deviations from normal respiration and to quantitatively define the time to symptom onset and disease duration. A 0.85 rabbit intramuscular median lethal dose of BoNT/A elicited the most consistent and prolonged disease duration (mean = 11.8 days, relative standard deviation = 27.9%) that still enabled spontaneous recovery. Post-exposure treatment with antitoxin at various time points significantly shortened the disease duration, providing a proof of concept that the new model is adequate for evaluating novel therapeutics for botulism.

Botulinum neurotoxins (BoNT) are some of the most toxic proteins known and can induce respiratory failure requiring long-term intensive care. Treatment of botulism includes the administration of antitoxins. Monoclonal antibodies (mAbs) hold considerable promise as BoNT therapeutics and prophylactics, due to their potency and safety. A three-mAb combination has been developed that specifically neutralizes BoNT serotype A (BoNT/A), and a separate three mAb combination has been developed that specifically neutralizes BoNT serotype B (BoNT/B). A six mAb cocktail, designated G03-52-01, has been developed that combines the anti-BoNT/A and anti-BoNT/B mAbs. The pharmacokinetics and neutralizing antibody concentration (NAC) of G03-52-01 has been determined in guinea pigs, and these parameters were correlated with protection against an inhalation challenge of BoNT/A1 or BoNT/B1. Previously, it was shown that each antibody demonstrated a dose-dependent mAb serum concentration and reached maximum circulating concentrations within 48 h after intramuscular (IM) or intraperitoneal (IP) injection and that a single IM injection of G03-52-01 administered 48 h pre-exposure protected guinea pigs against an inhalation challenge of up to 93 LD 50 s of BoNT/A1 and 116 LD 50 s of BoNT/B1. The data presented here advance our understanding of the relationship of the neutralizing NAC to the measured circulating antibody concentration and provide additional support that a single IM or intravenous (IV) administration of G03-52-01 will provide pre-exposure prophylaxis against botulism from an aerosol exposure of BoNT/A and BoNT/B.

Human serum albumin (HSA) has been widely used as a pharmaceutical excipient in Botulinum toxin serotype A (BoNT/A) products that are indicated for use in therapeutics and cosmetics. However, HSA as a human-derived material has some concerns, such as the potential risk of transmission of infectious agents, an insufficient supply, and difficulty in maintaining a certain quality. For those reasons, newly developed BoNT/A products (CORETOX®, Medytox, Inc., Republic of Korea) contained polysorbate 20, a non-human-derived excipient, to replace the HSA. However, most safety studies of polysorbate 20 have been conducted with non-invasive routes of administration, and thus there are a few studies on the safety of polysorbate 20 when administered intramuscularly. To secure the in vivo safety profile of polysorbate 20, a four-week repeated intramuscular dose toxicity study (0.02, 0.1, and 0.4 mg/kg, one injection every two weeks for a total of three injections) was conducted in 66 Sprague-Dawley (SD) rats. An intradermal irritation study was further conducted with 18 New Zealand White (NZW) rabbits. The toxicological evaluation of HSA (0.06 and 0.12 mg/kg) was also carried out as a comparative substance. Systemic and local toxicities were not observed in any of the SD rats or NZW rabbits based on clinical signs, body weight, hematology, clinical biochemistry, macroscopic findings on necropsy, histopathology of the injection site, and allergic reactions. The current study suggested that intramuscular administration of polysorbate 20 was considered to be safe at a level similar to that of HSA, which has an in vivo safety profile accumulated over the years. This provided the basis for the in vivo safety profile of polysorbate 20 administered intramuscularly and the scientific reliability of the use of polysorbate 20 as an alternative to HSA, which is used as an excipient for various pharmaceuticals in terms of its safety., Competing Interests: The authors have declared that no competing interests exist. Furthermore, the authors [JK, SK, MSP, CHR, GHY, WhK] employed by a commercial company, Medytox Inc., confirm that this commercial affiliation does not alter their adherence to PLOS ONE policies on sharing data and materials.

Botulinum neurotoxins (BoNTs) are the most poisonous substances in nature. Currently, the only therapy for botulism is antitoxin. This therapy suffers from several limitations and hence new therapeutic strategies are desired. One of the limitations in discovering BoNT inhibitors is the absence of an in vitro assay that correlates with toxin neutralization in vivo. In this work, a high-throughput screening assay for receptor-binding inhibitors against BoNT/A was developed. The assay is composed of two chimeric proteins: a receptor-simulating protein, consisting of the fourth luminal loop of synaptic vesicle protein 2C fused to glutathione-S-transferase, and a toxin-simulating protein, consisting of the receptor-binding domain of BoNT/A fused to beta-galactosidase. The assay was applied to screen the LOPAC1280 compound library. Seven selected compounds were evaluated in mice exposed to a lethal dose of BoNT/A. The compound aurintricarboxylic acid (ATA) conferred 92% protection, whereas significant delayed time to death ( p < 0.005) was observed for three additional compounds. Remarkably, ATA was also fully protective in mice challenged with a lethal dose of BoNT/E, which also uses the SV2 receptor. This study demonstrates that receptor-binding inhibitors have the potential to serve as next generation therapeutics for botulism, and therefore the assay developed may facilitate discovery of new anti-BoNT countermeasures.

Antitoxin is currently the only approved therapy for botulinum intoxications. The efficacy of antitoxin preparations is evaluated in animals. However, while in practice antitoxin is administered to patients only after symptom onset, in most animal studies, it is tested in relation to time postintoxication. This may be attributed to difficulties in quantitating early botulism symptoms in animals. In the current study, a novel system based on high-resolution monitoring of mouse activity on a running wheel was developed to allow evaluation of postsymptom antitoxin efficacy. The system enables automatic and remote monitoring of 48 mice simultaneously. Based on the nocturnal activity patterns of individual naive mice, two criteria were defined as the onset of symptoms. Postsymptom treatment with a human-normalized dose of antitoxin was fully protective in mice exposed to 4 50% lethal doses (LD 50 s) of botulinum neurotoxin serotype A (BoNT/A) and BoNT/B. Moreover, for the first time, a high protection rate was obtained in mice treated postsymptomatically, following a challenge with BoNT/E, the fastest-acting BoNT. The running wheel system was further modified to develop a mouse model for the evaluation of next-generation therapeutics for progressive botulism at time points where antitoxin is not effective. Exposure of mice to 0.3 LD 50 of BoNT/A resulted in long-lasting paralysis and a reduction in running activity for 16 to 18 days. Antitoxin treatment was no longer effective when administered 72 h postintoxication, defining the time window to evaluate next-generation therapeutics. Altogether, the running wheel systems presented herein offer quantitative means to evaluate the efficacy of current and future antibotulinum drugs.

Botulism is a rare, life-threatening paralytic disease caused by Clostridium botulinum neurotoxin (BoNT). Available treatments, including an equine antitoxin and human immune globulin, are given postexposure and challenging to produce and administer. NTM-1632 is an equimolar mixture of 3 human IgG monoclonal antibodies, B1, B2, and B3, targeting BoNT serotype B (BoNT/B). This first-in-human study assessed the safety, tolerability, pharmacokinetics (PK), and immunogenicity of NTM-1632. This double-blind, single-center, placebo-controlled dose escalation study randomized 3 cohorts of healthy volunteers to receive a single intravenous dose of NTM-1632 (0.033, 0.165, or 0.330 mg/kg) or saline placebo. Safety monitoring included physical examinations, clinical laboratory studies, and vital signs. Blood sampling was performed at prespecified time points for PK and immunogenicity analyses. Twenty-four subjects received study product (18 NTM-1632; 6 placebo), and no deaths or serious adverse events were reported. Adverse events in the NTM-1632 groups were generally mild and similar in frequency and severity to the placebo group, and no safety signal was identified. NTM-1632 has a favorable PK profile with a half-life of >20 days for the 0.330-mg/kg dose and an approximately linear relationship with respect to maximum concentration and area under the concentration-time curve (AUC 0→ t ). NTM-1632 demonstrated low immunogenicity with only a few treatment-emergent antidrug antibody responses in the low and middle dosing groups and none at the highest dose. NTM-1632 is well tolerated at the administered doses. The favorable safety, PK, and immunogenicity profile of NTM-1632 supports further clinical development as a treatment for BoNT/B intoxication and postexposure prophylaxis. (This study has been registered at ClinicalTrials.gov under identifier NCT02779140.).

Background: Small bowel obstruction is one of the leading reasons for accessing to the Emergency Department. Food poisoning from Clostridium botulinum has emerged as a very rare potential cause of small bowel obstruction. The relevance of this case report regards the subtle onset of pathognomonic neurological symptoms, which can delay diagnosis and subsequent life-saving treatment., Case Presentation: A 24-year-old man came to our Emergency Department complaining of abdominal pain, fever and sporadic self-limiting episodes of diplopia, starting 4 days earlier. Clinical presentation and radiological imaging suggested a case of small bowel obstruction. Non-operative management was adopted, which was followed by worsening of neurological signs. On specifically questioning the patient, we discovered that his parents had experienced similar, but milder symptoms. The patient also recalled eating home-made preserves some days earlier. A clinical diagnosis of foodborne botulism was established and antitoxin was promptly administered with rapid clinical resolution., Conclusions: Though very rare, botulism can mimic small bowel obstruction, and could be associated with a rapid clinical deterioration if misdiagnosed. An accurate family history, frequent clinical reassessments and involvement of different specialists can guide to identify this unexpected diagnosis.

Botulism is caused by a potent neurotoxin that blocks neuromuscular transmission, resulting in death by asphyxiation. Currently, the therapeutic options are limited and there is no antidote. Here, we harness the structural and trafficking properties of an atoxic derivative of botulinum neurotoxin (BoNT) to transport a function-blocking single-domain antibody into the neuronal cytosol where it can inhibit BoNT serotype A (BoNT/A1) molecular toxicity. Post-symptomatic treatment relieved toxic signs of botulism and rescued mice, guinea pigs, and nonhuman primates after lethal BoNT/A1 challenge. These data demonstrate that atoxic BoNT derivatives can be harnessed to deliver therapeutic protein moieties to the neuronal cytoplasm where they bind and neutralize intracellular targets in experimental models. The generalizability of this platform might enable delivery of antibodies and other protein-based therapeutics to previously inaccessible intraneuronal targets., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Efficient penetration of cell membranes and specific targeting of a cell type represent major challenges for developing therapeutics toward intracellular targets. One example facing these hurdles is to develop post-exposure treatment for botulinum neurotoxins (BoNTs), a group of bacterial toxins (BoNT/A to BoNT/G) that are major potential bioterrorism agents. BoNTs enter motor neurons, block neurotransmitter release, and cause a paralytic disease botulism. Members of BoNTs such as BoNT/A exhibit extremely long half-life within neurons, resulting in persistent paralysis for months, yet there are no therapeutics that can inhibit BoNTs once they enter neurons. Here, we developed a chimeric toxin-based delivery platform by fusing the receptor-binding domain of a BoNT, which targets neurons, with the membrane translocation domain and inactivated protease domain of the recently discovered BoNT-like toxin BoNT/X, which can deliver cargoes across endosomal membranes into the cytosol. A therapeutic protein was then created by fusing a single-domain antibody (nanobody) against BoNT/A with the delivery platform. In vitro characterization demonstrated that nanobodies were delivered into cultured neurons and neutralized BoNT/A in neurons. Administration of this protein in mice shortened duration of local muscle paralysis, restoring muscle function within hours, and rescued mice from systemic toxicity of lethal doses of BoNT/A. Fusion of two nanobodies, one against BoNT/A and the other against BoNT/B, created a multivalent therapeutic protein able to neutralize both BoNT/A and BoNT/B in mice. These studies provide an effective post-exposure treatment for botulism and establish a platform for intracellular delivery of therapeutics targeting cytosolic proteins and processes., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Botulinum neurotoxins (BoNT) are extremely potent and can induce respiratory failure, requiring long-term intensive care to prevent death. Recombinant monoclonal antibodies (mAbs) hold considerable promise as BoNT therapeutics and prophylactics. In contrast, equine antitoxin cannot be used prophylactically and has a short half-life. Two three-mAb combinations are in development that specifically neutralize BoNT serotype A (BoNT/A) and B (BoNT/B). The three-mAb combinations addressing a single serotype provided pre-exposure prophylaxis in the guinea pig inhalation model. A lyophilized co-formulation of six mAbs, designated G03-52-01, that addresses both A and B serotypes is in development. Here, we investigated the efficacy of G03-52-01 to protect guinea pigs against an aerosol exposure challenge of BoNT/A1 or BoNT/B1. Previously, it was found that each antibody demonstrated a dose-dependent exposure and reached maximum circulating concentrations within 48 h after intramuscular (IM) or intravenous (IV) injection. Here we show that G03-52-01, in a single IM injection of G03-52-01 administered 48 h pre-exposure, protected guinea pigs against an aerosol challenge of up to 238 LD 50 s of BoNT/A1 and 191 LD 50 s of BoNT/B1. These data suggest that a single IM administration of G03-52-01 provides pre-exposure prophylaxis against botulism from an aerosol exposure of BoNT/A1 or BoNT/B1.

Botulinum toxin is nowadays approved as an effective medication for various neurological disorders. The extreme toxicity of this toxin-inducing botulism, a severe lethal muscle-paralyzing illness, has been well known since the seminal works of the end of the 19th century. Because of this toxicity, botulinum toxin was one of the first agents to be considered for use as a biological weapon. The Second World War was a crucial period for the first attempts to weaponize this toxin even if many unknown factors about botulinum toxin still existed at the outbreak of the war. Using documents from the British National Archives and other published sources, we discuss the main points of the attempts to weaponize this toxin in German and Allied armies. During WW2, Allied intelligence services regularly reported a major German threat related to the potential use of botulinum toxin as a biological weapon, especially during the preparation of Operation Overlord, the Allied invasion to liberate Europe. All these reports would ultimately prove to be inaccurate: botulinum toxin was not part of the German military arsenal even if some German scientists tried to use the results of the French pre-war military research. Misinformation spread by intelligence services stimulated military research at Porton Down facilities in England and at Camp Detrick in the USA. These studies led to a succession of failures and myths about the weaponization of botulinum toxin. Nevertheless, major progress (purification, toxoid) arose from the military research, providing useful data for the first steps of the therapeutic use of botulinum toxin in the post-war years., (© 2021 S. Karger AG, Basel.)

Botulinum neurotoxins (BoNTs), produced by Clostridia and other bacteria, are the most potent toxins known. Their cleavage of the soluble N-ethylmaleimide-sensitive factor activating protein receptor (SNARE) proteins in neurons prevents the release of neurotransmitters, thus resulting in the muscle paralysis that is characteristic of botulism. This mechanism of action has been exploited for a variety of therapeutic and cosmetic applications of BoNTs. This chapter provides an overview of the native BoNTs, including the classical serotypes and their clinical use, mosaic BoNTs, and novel BoNTs that have been recently identified in clostridial and non-clostridial strains. In addition, the modular structure of native BoNTs, which are composed of a light chain and a heavy chain, is amenable to a multitude of novel fusions and mutations using molecular biology techniques. These novel recombinant BoNTs have been used or are being developed to further characterize the biology of toxins, to assist in vaccine production, to serve as delivery vehicles to neurons, and to be utilized as novel therapeutics for both neuronal and non-neuronal cells.

Infant botulism is a progressive process described as starting with descending weakness, facial palsies and constipation. Loss of bulbar reflexes and flaccid paralysis are common in infants less than 6 months old who have infant botulism. Clostridium botulinum , the bacteria that produce the toxin that causes this condition, are ubiquitous in the United States including Hawai'i, but infant botulism is rarely reported here. This report describes 2 cases of infant botulism with atypical initial presentations diagnosed on O'ahu, Hawai'i. Patient A is a 3-month-old male who presented with altered mental status, including inconsolability, who progressed to loss of gag reflex and constipation. Due to early concern for meningitis, Patient A was treated with antibiotics, however further evaluation led to eventual positive testing for botulinum B toxin. Patient B is a 2-month-old female who presented with somnolence and fever after immunizations and progressed to respiratory failure and apparent dehydration. Because she presented shortly after receiving immunizations, metabolic disorders were strongly considered as a potential cause of symptoms, but Patient B had normal metabolic evaluation and eventually tested positive for botulinum A toxin. Altered mental status and fever are unusual presentations for infant botulism. Infant botulism should be considered in infants with altered mental status when the course of illness includes the development of constipation and weakness, and evaluations are not suggestive of alternative causes, including infection, metabolic diseases, and spinal muscular atrophy. Early consideration and treatment of infant botulism should be considered for infants presenting with altered mental status who develop neuromuscular weakness. The Infant Botulism Treatment and Prevention Program (www.infantbotulism.org) should be contacted early for assistance with diagnosis and treatment., Competing Interests: None of the authors identify a conflict of interest., (©Copyright 2020 by University Health Partners of Hawai‘i (UHP Hawai‘i).)

Background: Botulism is a rare, life-threatening paralytic illness. Botulism Antitoxin Heptavalent (A,B,C,D,E,F,G)-(Equine) (BAT) manufactured by Emergent BioSolutions Canada Inc is an equine-derived heptavalent botulinum antitoxin product indicated for the treatment of symptomatic botulism following documented or suspected exposure to botulinum neurotoxin serotypes A-G in adults and pediatric patients. BAT product was US-licensed in 2013., Methods: In the United States, from October 2014 through July 2017, safety and clinical outcomes data were collected under a registry for patients treated with BAT product., Results: Registry patients had a median age of 51 years (range, 32 days to 92 years). Among 162 patients, 7 (4.3%) experienced BAT product-related serious adverse events, including 1 (0.6%) report each of pneumonia, pneumonia aspiration, ventricular tachycardia, upper gastrointestinal hemorrhage, anaphylactic reaction, acute kidney injury, and acute myocardial infarction. Thirty-one (19.1%) patients had 41 BAT product-related adverse events. Six (3.7%) deaths were reported in the registry. All deaths were attributed to the underlying illness and were assessed as unlikely related to BAT product. Among 113 (69.8%) patients with a final diagnosis of botulism, those treated early (≤2 days) spent fewer days in the hospital (5 vs 15.5 days), in the intensive care unit (ICU) (4 vs 12 days), and on mechanical ventilation (6 vs 14.5 days) than those treated late (>2 days), respectively., Conclusions: BAT product was well tolerated in patients. Treatment with BAT product at ≤2 days of symptom onset was associated with shorter hospital and ICU stays, and shorter duration and need for mechanical ventilation, showing clinical benefit associated with early treatment., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

A 44-year-old man with a background of heroin injection drug use was referred to the ear, nose and throat team with a sore throat and dysphagia. He was treated with intravenous antibiotics and steroids for suspected uvulitis. He developed progressive bulbar weakness and symmetrical descending weakness of the upper extremities over a 12-hour period and was intubated prior to transfer to the intensive care unit.Botulinum heptavalent antitoxin was administered, and subsequent PCR assay confirmed Clostridium botulinum neurotoxin B from his most recent injection site. He was found unconscious on the ward 3 days following extubation. Postmortem confirmed he died from heroin intoxication.This case highlights the importance of considering wound botulism in injection drug users presenting with unexplained weakness, particularly of the lower cranial nerves. Botulism is not characteristically associated with signs of localised or systemic infection in contrary to other bacterial complications of injection drug use., Competing Interests: Competing interests: None declared., (© BMJ Publishing Group Limited 2020. No commercial re-use. See rights and permissions. Published by BMJ.)

Botulinum neurotoxins (BoNTs) are potent neuroparalytic toxins that cause mortality through respiratory paralysis. The approved medical countermeasure for BoNT poisoning is infusion of antitoxin immunoglobulins. However, antitoxins have poor therapeutic efficacy in symptomatic patients; thus, there is an urgent need for treatments that reduce the need for artificial ventilation. We report that the US Food and Drug Administration-approved potassium channel blocker 3,4-diaminopyridine (3,4-DAP) reverses respiratory depression and neuromuscular weakness in murine models of acute and chronic botulism. In ex vivo studies, 3,4-DAP restored end-plate potentials and twitch contractions of diaphragms isolated from mice at terminal stages of BoNT serotype A (BoNT/A) botulism. In vivo, human-equivalent doses of 3,4-DAP reversed signs of severe respiratory depression and restored mobility in BoNT/A-intoxicated mice at terminal stages of respiratory collapse. Multiple-dosing administration of 3,4-DAP improved respiration and extended survival at up to 5 LD50 BoNT/A. Finally, 3,4-DAP reduced gastrocnemius muscle paralysis and reversed respiratory depression in sublethal models of serotype A-, B-, and E-induced botulism. These findings make a compelling argument for repurposing 3,4-DAP to symptomatically treat symptoms of muscle paralysis caused by botulism, independent of serotype. Furthermore, they suggest that 3,4-DAP is effective for a range of botulism symptoms at clinically relevant time points.

Botulinum neurotoxin (BoNT) is widely used for the treatment of spasticity, focal dystonia, chronic migraine, facial hemispasm, and facial aesthetic treatments. Generally, treatment with botulinum toxin is a safe procedure when conducted by clinicians with expertise, and local side effects are rare and transient. However, occasionally adverse effects can occur due to the spread of the drug to nontargeted muscles and organs, producing dry mouth, fatigue, and flu-like symptoms, up to signs of systemic botulism, which appears to be more frequent in children treated for spasticity than in adults. In silico 3D-QSAR and molecular docking studies were performed to build a structure-based model on selected potent known botulinum neurotoxin type A inhibitors; this was used to screen the US Food and Drug Administration (FDA) database. Thirty molecules were identified as possible light-chain BoNT/A inhibitors. In this study, we applied a well-established ligand- and structure-based methodology for the identification of hit compounds among a database of FDA-approved drugs. The identification of budesonide, protirelin, and ciclesonide followed by other compounds can be considered a starting point for investigations of selected compounds that could bypass much of the time and costs involved in the drug approval process., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Botulism caused by toxins of Clostridium botulinum and other neurotoxic clostridia is a rare but life-threatening disease with neurological symptoms. Food-borne botulism (food poisoning) is the most common type worldwide; rarely, wound botulism, infant botulism, or botulism of unknown etiology may also occur. Botulism is a very rare disease in the Czech Republic as well, with only nine cases reported since 2008 (EPIDAT). These were mostly sporadic cases of food-borne botulism except a small family outbreak with three cases due to the consumption of homemade pork pate (2013). This outbreak prompted the creation of a national standby supply of life-saving anti-infective drugs in Prague. This article reviews the etiology, epidemiology, clinical manifestations, diagnosis, and therapy of botulism.

Purpose of Review: This article reviews the pathophysiology, epidemiology, clinical presentation, diagnosis, and treatment of Lambert-Eaton myasthenic syndrome (LEMS) and of botulism, and immune-related myasthenia gravis (MG) occurring in the context of immune checkpoint inhibitor therapy for cancer., Recent Findings: The suspicion that LEMS is rare but also likely underdiagnosed is supported by recent epidemiologic data. A validated, LEMS-specific scale now exists to assess and monitor disease, and symptomatic and immunomodulatory treatments are available. As presynaptic disorders of neuromuscular transmission, LEMS and botulism share electrodiagnostic abnormalities but have important distinguishing features. Knowledge of the clinical features of botulism is needed, particularly with continued cases of infant botulism, the opioid epidemic increasing the incidence of wound botulism, and medical use of botulinum toxin, which may cause iatrogenic botulism. Foodborne botulism remains rare. Prompt recognition of botulism and administration of antitoxin can improve outcomes. MG may be exacerbated or may present de novo in the context of immune activation from immune checkpoint inhibitor therapies for cancer. Immune-related MG commonly overlaps with myositis and myocarditis. Corticosteroids typically result in improvement. However, immune-related MG can be more fulminant than its idiopathic counterpart and may cause permanent disability or death., Summary: The diagnosis of LEMS, botulism, or immune-related MG can generally be made from the patient's history, supplemented with directed questions, a physical examination designed to demonstrate abnormalities, and laboratory and electrodiagnostic testing. Early diagnosis and carefully selected treatment not only improve outcomes of the neuromuscular disease but can affect the prognosis of underlying malignancy, when present.

Background: Botulism is a rare, serious, and sometimes fatal paralytic illness caused by exposure to neurotoxins produced by Clostridium botulinum bacteria. Patients with documented or suspected exposure to botulinum toxin serotypes A-G can be treated with BAT® [Botulism Antitoxin Heptavalent (A, B, C, D, E, F, G)-(Equine)] product, which was approved in 2013 in the United States (US). Patients with botulism have demonstrated greater clinical benefit with early BAT product treatment (≤2 days from symptom onset) versus late treatment (>2 days)., Objective: Economic outcomes associated with improved clinical outcome benefits of BAT product treatment have not yet been reported. This ad hoc analysis aimed to estimate and compare costs associated with hospitalization, intensive care unit stay, and mechanical ventilation for patients with botulism administered BAT product treatment early or late., Methods: Clinical outcomes data for early and late BAT product treatment were obtained from a patient registry conducted between October 2014 and July 2017. Total per patient mean daily costs were estimated based on information from published literature. Total population costs per group were calculated by multiplying estimated mean cost per patient by the average annual number of non-infant botulism cases in the US., Results: Mean per patient costs were 2.5 times lower for patients treated with BAT product early versus late. On average in the US, early BAT product treatment could save greater than $3.9 million per year versus late treatment., Conclusion: Substantial economic savings can be achieved with early BAT product treatment. The findings support the recommendation for public health authorities to ensure antitoxin treatment is readily available in sufficient quantities to manage botulism cases, including sporadic outbreaks and potential mass exposure biological attacks., Competing Interests: DMA and JSR are employees of Emergent BioSolutions Canada Inc. and VRK is an employee of Emergent BioSolutions Inc. DMA and VRK hold stock. DLA and SPB are employees of IQVIA, which received funding from Emergent BioSolutions Inc. to conduct this analysis. EK was an employee of IQVIA at the time this study was conducted. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

A large number of bacterial toxins consist of active and cell binding protomers linked by an interchain disulfide bridge. The largest family of such disulfide-bridged exotoxins is that of the clostridial neurotoxins that consist of two chains and comprise the tetanus neurotoxins causing tetanus and the botulinum neurotoxins causing botulism. Reduction of the interchain disulfide abolishes toxicity, and we discuss the experiments that revealed the role of this structural element in neuronal intoxication. The redox couple thioredoxin reductase-thioredoxin (TrxR-Trx) was identified as the responsible for reduction of this disulfide occurring on the cytosolic surface of synaptic vesicles. We then discuss the very relevant finding that drugs that inhibit TrxR-Trx also prevent botulism. On this basis, we propose that ebselen and PX-12, two TrxR-Trx specific drugs previously used in clinical trials in humans, satisfy all the requirements for clinical tests aiming at evaluating their capacity to effectively counteract human and animal botulism arising from intestinal toxaemias such as infant botulism., (© 2019 The Authors Cellular Microbiology Published by John Wiley & Sons Ltd.)

Botulinum neurotoxin serotype A (BoNT/A), marketed commercially as Botox, is the most toxic substance known to man with an estimated intravenous lethal dose (LD 50 ) of 1-2 ng/kg in humans. Despite its widespread use in cosmetic and medicinal applications, no postexposure therapeutics are available for the reversal of intoxication in the event of medical malpractice or bioterrorism. Accordingly, the Centers for Disease Control and Prevention categorizes BoNT/A as a Category A pathogen, posing the highest risk to national security and public health as a result of the ease with which BoNT/A can be weaponized and disseminated. BoNT/A-mediated lethality results from neurons impeded from releasing acetylcholine, which ultimately causes muscle paralysis and possible death by asphyxiation with the loss of diaphragm function. Currently, the only available respite for BoNT/A poisoning is antibody-based therapy; however, this intervention is only effective within 12-24 h postexposure. Small molecule therapeutics remain the only opportunity to reverse BoNT/A intoxication after neuronal poisoning and are urgently needed. Nevertheless, no small molecule BoNT/A inhibitors have reached the clinic or even advanced to clinical trials. This Account highlights the accomplishments and existing challenges facing BoNT/A drug discovery today. Using the comprehensive body of work from our laboratory, we illustrate our nearly two-decade endeavor to discover a clinically relevant BoNT/A inhibitor. Specifically, a discussion on the identification and characterization of new chemical leads, the development of in vitro and in vivo assays, and pertinent discoveries in BoNT/A structural biology related to small molecule inhibition is presented. Lead discovery efforts in our laboratory have leveraged both in vitro high-throughput screening and rational design, and an array of mechanistic strategies for inhibiting BoNT/A has been discovered, including noncovalent inhibition, metal-binding active site inhibition, covalent inhibition, and α- and β-exosite inhibition. We contrast the strengths and weaknesses of each of these mechanistic strategies and propose the most favorable approach for success. Finally, we discuss multiple serendipitous discoveries of antibotulism small molecules with alternative mechanisms of action. Remaining challenges facing clinically relevant BoNT/A inhibition are presented and analyzed, including the current inability to reconcile toxin half-life (months to greater than one year) in neurons with in vivo pharmaceutical lifetimes and reoccurring inconsistencies between in vitro , cellular, and in vivo translation. Our Account of BoNT/A chemical research emphasizes the present accomplishments and critically analyzes the remaining obstacles for drug discovery. Importantly, we call for an increased focus on the discovery of safe and effective covalent inhibitors of BoNT/A that compete with the inherent half-life of the toxin.

Background: Botulism is a disease caused by neurogenic toxins that block acetylcholine release, resulting in potentially life threatening neuroparalysis. Seven distinct serotypes of botulinum neurotoxins (BoNTs) have been described and are found in nature world-wide. This, combined with ease of production, make BoNTs a significant bioweapon threat. An essential countermeasure to this threat is an antitoxin to remove circulating toxin. An antitoxin, tradename BAT (Botulism Antitoxin Heptavalent (A, B, C, D, E, F, G)-(Equine)), has been developed and its efficacy evaluated against all seven serotypes in guinea pigs., Methods and Findings: Studies were conducted to establish the lethal dose and clinical course of intoxication for all seven toxins, and post-exposure prophylactic efficacy of BAT product. Animals were monitored for signs of intoxication and mortality for 14 days. Guinea pig intramuscular LD50s (GPIMLD50) for all BoNTs ranged from 2.0 (serotype C) to 73.2 (serotype E) of mouse intraperitoneal LD50 units. A dose of 4x GPIMLD50 was identified as the appropriate toxin dose for use in subsequent efficacy and post-exposure prophylaxis studies. The main clinical signs observed included hind limb paralysis, weak limb, change in breathing rate/pattern, and forced abdominal respiration. Mean time to onset of clinical signs ranged from 12 hours (serotype E) to 39 hours (serotype G). Twelve hours post-intoxication was selected as the appropriate time point for intervention for all serotypes apart from E where 6 hours was selected because of the rapid onset and progression of clinical signs. Post-exposure treatment with BAT product resulted in a significantly (p<0.0001) higher survival at >0.008 scaled human dose for serotypes A, B, C, F and G, at >0.2x for serotype D and >0.04x for serotype E., Conclusions: These studies confirm the efficacy of BAT as a post-exposure prophylactic therapy against all seven known BoNT serotypes., Competing Interests: A. Emanuel and T. Takla were employees of Emergent BioSolutions Canada Inc. at the time of the study. Doug Barker, Hongyu Qiu, and Shantha Kodihalli are current employees of Emergent BioSolutions Canada Inc., who developed and manufactured the BAT and placebo employed in these studies. Karen Gillum, Nancy Neimuth are employees of Battelle Biomedical Research Center and claim no conflict of interest. The specific roles of these authors are articulated in the ‘author contributions’ section. This does not alter the authors’ adherence to PLOS ONE policies on sharing data and materials. Emergent BioSolutions, Protected by Emergent BioSolutions, BAT, and any and all Emergent BioSolutions Inc. brand, product, service and feature names, logos and slogans are trademarks or registered trademarks of Emergent BioSolutions Inc. or its subsidiaries in the United States or other countries. All other brand, product, service and feature names or trademarks are the property of their respective owners.

Background: Clostridium botulinum causes botulism, a serious paralytic illness that results from the ingestion of a botulinum toxin. Because silver nanoparticle products exhibit strong antimicrobial activity, applications for silver nanoparticles in healthcare have expanded. Therefore, the objective of the current study was to assess a therapeutic strategy for the treatment of botulism toxicity using silver nanoparticles., Methods: A preliminary test was conducted using doses that produce illness in laboratory animals to determine the absolute lethal dose (LD100) of botulinum toxin type A (BoNT/A) in mice. Next, the test animals were divided into six groups containing six mice each. Groups I, II and III were the negative control (botulinum toxin only), positive control-1 (nano-silver only) and positive control-2 (no treatment), respectively. The remaining groups were allocated to the toxin that was supplemented with three nano-silver treatments., Results: The mortality rates of mice caused by BoNT/A significantly reduced in the treatment groups with different doses and injection intervals of nano-silver when compared to the negative control group. BoNT/A toxicity induced by intraperitoneal injection of the toxin of Clostridium botulinum causes rapid death while when coupled with nano-osilver results in delayed death in mice., Conclusion: These results, while open to future improvement, represent a preliminary step towards the satisfactory control of BoNT/A with the use of silver nanoparticles for human protection against this bioterrorism threat. Further study in this area can elucidate the underlying mechanism for detoxifying BoNT/A by silver nanoparticles., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Infant botulism is an infectious intestinal toxemia that results from colonization of the infant large bowel by Clostridium botulinum (or rarely, by neurotoxigenic Clostridium baratii or Clostridium butyricum ), with subsequent intraintestinal production and absorption of botulinum neurotoxin that then produces flaccid paralysis. The disease is often initially misdiagnosed as suspected sepsis or meningitis, diagnoses that require prompt empirical antimicrobial therapy. Antibiotics may also be needed to treat infectious complications of infant botulism, such as pneumonia or urinary tract infection. Clinical evidence suggests (see case report below) that broad-spectrum antibiotics that are eliminated by biliary excretion may cause progression of the patient's paralysis by lysing C. botulinum vegetative cells in the large bowel lumen, thereby increasing the amount of botulinum neurotoxin available for absorption. The purpose of this antimicrobial susceptibility study was to identify an antimicrobial agent with little or no activity against C. botulinum that could be used to treat infant botulism patients initially diagnosed with suspected sepsis or meningitis, or who acquired secondary infections, without lysing C. botulinum Testing of 12 antimicrobial agents indicated that almost all California infant botulism patient isolates are susceptible to most clinically utilized antibiotics and are also susceptible to newer antibiotics not previously tested against large numbers of C. botulinum patient isolates. No antibiotic with little or no activity against C. botulinum was identified. These findings reinforce the importance of promptly treating infant botulism patients with human botulism immune globulin (BIG-IV [BabyBIG])., (Copyright © 2018 American Society for Microbiology.)

Clostridia cause severe diseases. Tetanus is rare in Switzerland because of vaccine coverage and the application of guidelines for the management of contaminated wounds. Tetanus requires wound debridement and the administration of antibiotics and anti-tetanus immune. Besides gastroenteritis, infections due to C. perfringens most often require surgery, in addition to antibiotic treatment with penicillin and clindamycin. Botulism is a rare disease caused by a toxin produced by C. botulinum that causes flaccid paralysis. The clinical syndrome must be recognized early in order to administer the antitoxin and improve the prognosis. The other, rarer species of Clostridia require surgical and antibiotic management, but their prognosis remains poor., Competing Interests: Les auteurs n’ont aucun conflit d’intérêts à déclarer en relation avec cet article.

Botulinum neurotoxin (BoNT) serotypes A, B and E are responsible for most cases of human botulism. The only approved therapy for botulism is antitoxin treatment administered to patients after symptom onset. However, a recent meta-analysis of antitoxin efficacy in human botulism cases over the past century concluded that a statistically significant reduction in mortality is associated with the use of type E and type A antitoxin, but not with type B antitoxin. Animal models could be highly valuable in studying postsymptom antitoxin efficacy (PSAE). However, the few attempts to evaluate PSAE in animals relied on subjective observations and showed ∼50% protection. Recently, we developed a novel spirometry model for the quantitative evaluation of PSAE in rabbits and used it to demonstrate full protection against BoNT/E. In the current study, a comparative evaluation of PSAE in botulism types A and B was conducted using this quantitative respiratory model. A lethal dose of each toxin induced a comparable course of disease both in terms of time to symptoms (TTS, 41.9±1.3 and 40.6±1.1 h, respectively) and of time to death (TTD, 71.3±3.1 and 66.3±1.7 h, respectively). However, in accordance with the differential serotypic PSAE observed in humans, postsymptom antitoxin treatment was fully effective only in BoNT/A-intoxicated rabbits. This serotypic divergence was reflected by a positive and statistically significant correlation between TTS and TTD in BoNT/A-intoxicated rabbits (r=0.91, P =0.0006), but not in those intoxicated with BoNT/B (r=0.06, P =0.88). The rabbit spirometry system might be useful in the evaluation toolkit of botulism therapeutics, including those under development and intended to act when antitoxin is no longer effective., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

Competing Interests: Competing interests: None declared.

This study was conducted to analyze the clinical characteristics of and treatment strategies for botulism among patients receiving cosmetic injection of botulinum toxin (BoNT).A total of 86 botulism patients caused by cosmetic injection of BoNT were enrolled in our study. All of the patients were diagnosed according to their history of cosmetic BoNT injection, clinical symptoms and signs, and other auxiliary examinations (including those on renal and liver functions, blood index detection, and chest X-ray). All of the patients received comprehensive treatments and botulinum antitoxin serum injection.The main symptoms of botulism patients included headache, dizziness, insomnia, fatigue, blurred vision, eye opening difficulty, slurred speech, dysphagia, bucking, constipation, and anxiety. These clinical symptoms occurred 0∼36 days after BoNT injection, especially from 2nd to 6th day after the operation. Furthermore, the usage dose of BoNT was negatively related to latent period. Finally, patients all discharged from our hospital 1∼20 days after treatments, and their symptoms relieved or disappeared.Botulism is a severe side effect for BoNT injection. Injecting botulinum antitoxin serum may be an effective approach to improve clinical outcomes of botulism cases.