Publications by authors named "Michel R Popoff"

155 Publications

Characterization of a highly neutralizing single monoclonal antibody to botulinum neurotoxin type A.

FASEB J 2021 May;35(5):e21540

Institut Pasteur, Unité des Toxines Bactériennes, UMR CNRS 2001, Paris, France.

Compared to conventional antisera strategies, monoclonal antibodies (mAbs) represent an alternative and safer way to treat botulism, a fatal flaccid paralysis due to botulinum neurotoxins (BoNTs). In addition, mAbs offer the advantage to be produced in a reproducible manner. We previously identified a unique and potent mouse mAb (TA12) targeting BoNT/A1 with high affinity and neutralizing activity. In this study, we characterized the molecular basis of TA12 neutralization by combining Hydrogen/Deuterium eXchange Mass Spectrometry (HDX-MS) with site-directed mutagenesis and functional studies. We found that TA12 recognizes a conformational epitope located at the interface between the H and H subdomains of the BoNT/A1 receptor-binding domain (H ). The TA12-binding interface shares common structural features with the ciA-C2 VHH epitope and lies on the face opposite recognized by ciA-C2- and the CR1/CR2-neutralizing mAbs. The single substitution of N1006 was sufficient to affect TA12 binding to H confirming the position of the epitope. We further uncovered that the TA12 epitope overlaps with the BoNT/A1-binding site for both the neuronal cell surface receptor synaptic vesicle glycoprotein 2 isoform C (SV2C) and the GT1b ganglioside. Hence, TA12 potently blocks the entry of BoNT/A1 into neurons by interfering simultaneously with the binding of SV2C and to a lower extent GT1b. Our study reveals the unique neutralization mechanism of TA12 and emphasizes on the potential of using single mAbs for the treatment of botulism type A.
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http://dx.doi.org/10.1096/fj.202002492RDOI Listing
May 2021

Identification of a non-coding RNA and its putative involvement in the regulation of tetanus toxin synthesis in Clostridium tetani.

Sci Rep 2021 Feb 18;11(1):4157. Epub 2021 Feb 18.

Bacterial Toxins, Institut Pasteur, 25 rue du Dr Roux, 75724, Paris Cedex15, France.

Clostridium tetani produces the tetanus toxin (TeNT), one of the most powerful bacterial toxins known to humankind and responsible for tetanus. The regulation of toxin expression is complex and involves the alternative sigma factor TetR as well as other regulators. Here, a transcriptional analysis of the TeNT-encoding large plasmid of C. tetani identified a putative non-coding small RNA (sRNA), located in close vicinity of the 3' untranslated region of the tent gene. A northern blot experiment could identify a respective sRNA with a size of approx. 140 nucleotides. Sequence analysis showed that the sRNA contains a 14-nucleotide region that is complementary to a 5' located region of tent. In order to investigate the function of the sRNA, we applied a RNA interference approach targeting the sRNA in two C. tetani wild-type strains; the constructed antisense C. tetani strains showed an approx. threefold increase in both extracellular and total TeNT production compared to the respective wild-type strains. In addition, recombinant C. tetani strains were constructed that contained tent-locus harboring plasmids with and without the sRNA. However, the introduction of the tent-locus without the sRNA in a C. tetani strain lacking the wild-type TeNT-encoding large plasmid resulted in a lower TeNT production compared to the same strain with recombinant tent-locus with the sRNA. This suggests that the expression or the effect of the sRNA is modulated by the C. tetani genetic background, notably that of the wild-type TeNT-encoding large plasmid. In addition, some recombinant strains exhibited modulated growth patterns, characterized by premature bacterial cell lysis. Taken together, our data indicate that the sRNA acts as a negative regulator of TeNT synthesis, with a possible impact on the growth of C. tetani. We hypothesize that the role of this sRNA is to limit toxin levels in the exponential growth phase in order to prevent premature bacterial lysis.
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http://dx.doi.org/10.1038/s41598-021-83623-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7892561PMC
February 2021

Engineering Botulinum Neurotoxins for Enhanced Therapeutic Applications and Vaccine Development.

Toxins (Basel) 2020 Dec 22;13(1). Epub 2020 Dec 22.

Toxines Bacteriennes, Institut Pasteur, 75724 Paris, France.

Botulinum neurotoxins (BoNTs) show increasing therapeutic applications ranging from treatment of locally paralyzed muscles to cosmetic benefits. At first, in the 1970s, BoNT was used for the treatment of strabismus, however, nowadays, BoNT has multiple medical applications including the treatment of muscle hyperactivity such as strabismus, dystonia, movement disorders, hemifacial spasm, essential tremor, tics, cervical dystonia, cerebral palsy, as well as secretory disorders (hyperhidrosis, sialorrhea) and pain syndromes such as chronic migraine. This review summarizes current knowledge related to engineering of botulinum toxins, with particular emphasis on their potential therapeutic applications for pain management and for retargeting to non-neuronal tissues. Advances in molecular biology have resulted in generating modified BoNTs with the potential to act in a variety of disorders, however, in addition to the modifications of well characterized toxinotypes, the diversity of the wild type BoNT toxinotypes or subtypes, provides the basis for innovative BoNT-based therapeutics and research tools. This expanding BoNT superfamily forms the foundation for new toxins candidates in a wider range of therapeutic options.
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http://dx.doi.org/10.3390/toxins13010001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7821915PMC
December 2020

Toxemia in Human Naturally Acquired Botulism.

Toxins (Basel) 2020 11 13;12(11). Epub 2020 Nov 13.

Unité des Toxines Bactériennes, UMR CNRS 2001, Institut Pasteur, 75015 Paris, France.

Human botulism is a severe disease characterized by flaccid paralysis and inhibition of certain gland secretions, notably salivary secretions, caused by inhibition of neurotransmitter release. Naturally acquired botulism occurs in three main forms: food-borne botulism by ingestion of preformed botulinum neurotoxin (BoNT) in food, botulism by intestinal colonization (infant botulism and intestinal toxemia botulism in infants above one year and adults), and wound botulism. A rapid laboratory confirmation of botulism is required for the appropriate management of patients. Detection of BoNT in the patient's sera is the most direct way to address the diagnosis of botulism. Based on previous published reports, botulinum toxemia was identified in about 70% of food-borne and wound botulism cases, and only in about 28% of infant botulism cases, in which the diagnosis is mainly confirmed from stool sample investigation. The presence of BoNT in serum depends on the BoNT amount ingested with contaminated food or produced locally in the intestine or wound, and the timeframe between serum sampling and disease onset. BoNT levels in patient's sera are most frequently low, requiring a highly sensitive method of detection. Mouse bioassay is still the most used method of botulism identification from serum samples. However, in vitro methods based on BoNT endopeptidase activity with detection by mass spectrometry or immunoassay have been developed and depending on BoNT type, are more sensitive than the mouse bioassay. These new assays show high specificity for individual BoNT types and allow more accurate differentiation between positive toxin sera from botulism and autoimmune neuropathy patients.
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http://dx.doi.org/10.3390/toxins12110716DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7697460PMC
November 2020

Bacterial Toxins, Current Perspectives.

Authors:
Michel R Popoff

Toxins (Basel) 2020 09 4;12(9). Epub 2020 Sep 4.

Bacteries Anaerobies et Toxines, Institut Pasteur, 28 rue du Docteur Roux, 75724 Paris, France.

Toxins are the major pathogenicity factors produced by numerous bacteria involved in severe diseases in humans and animals. Certain pathogenic bacteria synthesize only one toxin which is responsible for all the symptoms and outcome of the disease. For example, botulinum toxins (BoNTs) and tetanus toxin (TeNT) are the unique causal factors of botulism and tetanus, respectively. Other bacteria attack the host organism by a set of multiple toxins which synergistically act to promote the disease. This is the case of and strains which secrete wide ranges of toxins such as pore-forming toxins, membrane phospholipid damaging toxins, and other cytotoxins and toxins interacting with the immune system involved in gangrene lesion generation.
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http://dx.doi.org/10.3390/toxins12090570DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7551839PMC
September 2020

Human Botulism in France, 1875-2016.

Toxins (Basel) 2020 05 21;12(5). Epub 2020 May 21.

Bacterial Toxins, Institut Pasteur, 75015 Paris, France.

Botulism is a rare but severe disease which is characterized by paralysis and inhibition of secretions. Only a few cases had been reported at the end of the 19th century in France. The disease was frequent during the second world war, and then the incidence decreased progressively. However, human botulism is still present in France with 10-25 cases every year. Food-borne botulism was the main form of botulism in France, whereas infant botulism (17 cases between 2004 and 2016) was rare, and wound and inhalational botulism were exceptional. Type B was the prevalent botulism type and was mainly due to consumption of home-made or small-scale preparations of cured ham and to a lesser extent other pork meat products. In the recent period (2000-2016), a wider diversity of botulism types from various food origin including industrial foods was reported. Severe cases of type A and F botulism as well as type E botulism were more frequent. Albeit rare, the severity of botulism justifies its continued surveillance and recommendations to food industry and consumers regarding food hygiene and preservation practices.
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http://dx.doi.org/10.3390/toxins12050338DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7291226PMC
May 2020

Central residues of the amphipathic β-hairpin loop control the properties of Clostridium perfringens epsilon-toxin channel.

Biochim Biophys Acta Biomembr 2020 09 22;1862(9):183364. Epub 2020 May 22.

Institut Pasteur, Bacterial Toxins, 28 rue du Dr Roux, 75724 Paris cedex 15, France. Electronic address:

Clostridium perfringens epsilon toxin (ETX) is a heptameric pore-forming toxin of the aerolysin toxin family. ETX is the most potent toxin of this toxin family and the third most potent bacterial toxin with high cytotoxic and lethal activities in animals. In addition, ETX shows a demyelinating activity in nervous tissue leading to devastating multifocal central nervous system white matter disease in ruminant animals. Pore formation in target cell membrane is most likely the initial critical step in ETX biological activity. Eight single to quadruple ETX mutants were generated by replacement of polar residues (serine, threonine, glutamine) in middle positions of the β-strands forming the β-barrel and facing the channel lumen with charged glutamic residues. Channel activity and ion selectivity were monitored in artificial lipid monolayer membranes and cytotoxicity was investigated in MDCK cells by the viability MTT test and propidium iodide entry. All the mutants formed channels with similar conductance in artificial lipid membranes and increasing cation selectivity for increasing number of mutations. Here, we show that residues in the central position of each β-strand of the amphipathic β-hairpin loop that forms the transmembrane pore, control the size and ion selectivity of the channel. While the highest cationic ETX mutants were not cytotoxic, no strict correlation was observed between ion selectivity and cytotoxicity.
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http://dx.doi.org/10.1016/j.bbamem.2020.183364DOI Listing
September 2020

Tetanus Toxin Synthesis is Under the Control of A Complex Network of Regulatory Genes in .

Toxins (Basel) 2020 05 15;12(5). Epub 2020 May 15.

Bactéries anaérobies et Toxines, Institut Pasteur, 75724 Paris, France.

produces a potent neurotoxin, the tetanus toxin (TeNT), which is responsible for an often-fatal neurological disease (tetanus) characterized by spastic paralysis. Prevention is efficiently acquired by vaccination with the TeNT toxoid, which is obtained by fermentation and subsequent purification and chemical inactivation. synthesizes TeNT in a regulated manner. Indeed, the TeNT gene () is mainly expressed in the late exponential and early stationary growth phases. The gene tetanus regulatory gene located immediately upstream of , encodes an alternative sigma factor which was previously identified as a positive regulator of . In addition, the genome of encodes more than 127 putative regulators, including 30 two-component systems (TCSs). Here, we investigated the impact of 12 regulators on TeNT synthesis which were selected based on their homology with related regulatory elements involved in toxin production in other clostridial species. Among nine TCSs tested, three of them impact TeNT production, including two positive regulators that indirectly stimulate and transcription. One negative regulator was identified that interacts with both and promoters. Two other TCSs showed a moderate effect: one binds to the promoter and weakly increases the extracellular TeNT level, and another one has a weak inverse effect. In addition, CodY (control of dciA (decoyinine induced operon) Y) but not Spo0A (sporulation stage 0) or the DNA repair protein Mfd (mutation frequency decline) positively controls TeNT synthesis by interacting with the promoter. Moreover, we found that inorganic phosphate and carbonate are among the environmental factors that control TeNT production. Our data show that TeNT synthesis is under the control of a complex network of regulators that are largely distinct from those involved in the control of toxin production in or .
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http://dx.doi.org/10.3390/toxins12050328DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7290440PMC
May 2020

Bacterial intracellularly active toxins: Membrane localisation of the active domain.

Cell Microbiol 2020 07 28;22(7):e13213. Epub 2020 May 28.

Bacterial Toxins, Institut Pasteur, Paris, France.

Numerous bacterial toxins exert their activity by inactivating or modulating a specific intracellular host target. For this purpose, these toxins have developed efficient strategies to overcome the different host cell defences including specific binding to cell surface, internalisation, passage through the endosome or plasma membrane, exploiting intracellular trafficking and addressing to intracellular targets. Several intracellularly active toxins deliver an active domain into the cytosol that interacts with a target localised to the inner face of the plasma membrane. Thus, the large clostridial glucosylating toxins (LCGTs) target Rho/Ras-GTPases, certain virulence factors of Gram negative bacteria, Rho-GTPases, while Pasteurella multocida toxin (PMT) targets trimeric G-proteins. Others such as botulinum neurotoxins and tetanus neurotoxin have their substrate on synaptic vesicle membrane. LCGTs, PMT, and certain virulence factors from Vibrio sp. show a particular structure constituted of a four-helix bundle membrane (4HBM) protruding from the catalytic site that specifically binds to the membrane phospholipids and then trap the catalytic domain at the proximity of the membrane anchored substrate. Structural and functional analysis indicate that the 4HBM tip of the Clostridium sordellii lethal toxin (TcsL) from the LCGT family contain two loops forming a cavity that mediates the binding to phospholipids and more specifically to phosphatidylserine.
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http://dx.doi.org/10.1111/cmi.13213DOI Listing
July 2020

Human peptide α-defensin-1 interferes with Clostridioides difficile toxins TcdA, TcdB, and CDT.

FASEB J 2020 05 19;34(5):6244-6261. Epub 2020 Mar 19.

Institute of Pharmacology and Toxicology, University of Ulm Medical Center, Ulm, Germany.

The human pathogenic bacterium Clostridioides difficile produces two exotoxins TcdA and TcdB, which inactivate Rho GTPases thereby causing C. difficile-associated diseases (CDAD) including life-threatening pseudomembranous colitis. Hypervirulent strains produce additionally the binary actin ADP-ribosylating toxin CDT. These strains are hallmarked by more severe forms of CDAD and increased frequency and severity. Once in the cytosol, the toxins act as enzymes resulting in the typical clinical symptoms. Therefore, targeting and inactivation of the released toxins are of peculiar interest. Prompted by earlier findings that human α-defensin-1 neutralizes TcdB, we investigated the effects of the defensin on all three C. difficile toxins. Inhibition of TcdA, TcdB, and CDT was demonstrated by analyzing toxin-induced changes in cell morphology, substrate modification, and decrease in transepithelial electrical resistance. Application of α-defensin-1 protected cells and human intestinal organoids from the cytotoxic effects of TcdA, TcdB, CDT, and their combination which is attributed to a direct interaction between the toxins and α-defensin-1. In mice, the application of α-defensin-1 reduced the TcdA-induced damage of intestinal loops in vivo. In conclusion, human α-defensin-1 is a specific and potent inhibitor of the C. difficile toxins and a promising agent to develop novel therapeutic options against C. difficile infections.
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http://dx.doi.org/10.1096/fj.201902816RDOI Listing
May 2020

Neuronal selectivity of botulinum neurotoxins.

Toxicon 2020 Apr 11;178:20-32. Epub 2020 Feb 11.

Bacterial Toxins, Institut Pasteur, Paris, France. Electronic address:

Botulinum neurotoxins (BoNTs) are highly potent toxins responsible for a severe disease, called botulism. They are also efficient therapeutic tools with an increasing number of indications ranging from neuromuscular dysfunction to hypersecretion syndrome, pain release, depression as well as cosmetic application. BoNTs are known to mainly target the motor-neurons terminals and to induce flaccid paralysis. BoNTs recognize a specific double receptor on neuronal cells consisting of gangliosides and synaptic vesicle protein, SV2 or synaptotagmin. Using cultured neuronal cells, BoNTs have been established blocking the release of a wide variety of neurotransmitters. However, BoNTs are more potent in motor-neurons than in the other neuronal cell types. In in vivo models, BoNT/A impairs the cholinergic neuronal transmission at the motor-neurons but also at neurons controlling secretions and smooth muscle neurons, and blocks several neuronal pathways including excitatory, inhibitory, and sensitive neurons. However, only a few reports investigated the neuronal selectivity of BoNTs in vivo. In the intestinal wall, BoNT/A and BoNT/B target mainly the cholinergic neurons and to a lower extent the other non-cholinergic neurons including serotonergic, glutamatergic, GABAergic, and VIP-neurons. The in vivo effects induced by BoNTs on the non-cholinergic neurons remain to be precisely investigated. We report here a literature review of the neuronal selectivity of BoNTs.
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http://dx.doi.org/10.1016/j.toxicon.2020.02.006DOI Listing
April 2020

Tetanus in animals.

Authors:
Michel R Popoff

J Vet Diagn Invest 2020 Mar 18;32(2):184-191. Epub 2020 Feb 18.

Bacterial Toxins, Institut Pasteur, Paris, Cedex, France.

Tetanus is a neurologic disease of humans and animals characterized by spastic paralysis. Tetanus is caused by tetanus toxin (TeNT) produced by , an environmental soilborne, gram-positive, sporulating bacterium. The disease most often results from wound contamination by soil containing spores. Horses, sheep, and humans are highly sensitive to TeNT, whereas cattle, dogs, and cats are more resistant. The diagnosis of tetanus is mainly based on the characteristic clinical signs. Identification of at the wound site is often difficult.
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http://dx.doi.org/10.1177/1040638720906814DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7081504PMC
March 2020

Report from the 26th Meeting on Toxinology, "Bioengineering of Toxins", Organized by the French Society of Toxinology (SFET) and Held in Paris, France, 4-5 December 2019.

Toxins (Basel) 2020 01 3;12(1). Epub 2020 Jan 3.

Service d'Ingénierie Moléculaire des Protéines (SIMOPRO), CEA de Saclay, Université Paris-Saclay,91191 Gif-sur-Yvette, France.

This 26th edition of the annual Meeting on Toxinology (RT26) of the SFET (http://sfet.asso.fr/international) was held at the Institut Pasteur of Paris on 4-5 December 2019 [...].
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http://dx.doi.org/10.3390/toxins12010031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7020397PMC
January 2020

Epsilon Toxin from Causes Inhibition of Potassium inward Rectifier (Kir) Channels in Oligodendrocytes.

Toxins (Basel) 2020 01 6;12(1). Epub 2020 Jan 6.

Université de Strasbourg, Institut des Neurosciences Cellulaires et Intégratives, Centre National de la Recherche Scientifique, 5 rue Blaise Pascal, F-67084 Strasbourg CEDEX, France.

Epsilon toxin (ETX), produced by types B and D, causes serious neurological disorders in animals. ETX can bind to the white matter of the brain and the oligodendrocytes, which are the cells forming the myelin sheath around neuron axons in the white matter of the central nervous system. After binding to oligodendrocytes, ETX causes demyelination in rat cerebellar slices. We further investigated the effects of ETX on cerebellar oligodendrocytes and found that ETX induced small transmembrane depolarization (by ~ +6.4 mV) in rat oligodendrocytes primary cultures. This was due to partial inhibition of the transmembrane inward rectifier potassium current (Kir). Of the two distinct types of Kir channel conductances (~25 pS and ~8.5 pS) recorded in rat oligodendrocytes, we found that ETX inhibited the large-conductance one. This inhibition did not require direct binding of ETX to a Kir channel. Most likely, the binding of ETX to its membrane receptor activates intracellular pathways that block the large conductance Kir channel activity in oligodendrocyte. Altogether, these findings and previous observations pinpoint oligodendrocytes as a major target for ETX. This supports the proposal that ETX might be a cause for Multiple Sclerosis, a disease characterized by myelin damage.
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http://dx.doi.org/10.3390/toxins12010036DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7020416PMC
January 2020

Public Health Risk Associated with Botulism as Foodborne Zoonoses.

Toxins (Basel) 2019 12 30;12(1). Epub 2019 Dec 30.

Institut Pasteur, Département de Microbiologie, Unité des Toxines Bactériennes, 25 rue du Dr Roux, 75015 Paris, France.

Botulism is a rare but severe neurological disease in man and animals that is caused by botulinum neurotoxins (BoNTs) produced by and atypical strains from other and non- species. BoNTs are divided into more than seven toxinotypes based on neutralization with specific corresponding antisera, and each toxinotype is subdivided into subtypes according to amino acid sequence variations. Animal species show variable sensitivity to the different BoNT toxinotypes. Thereby, naturally acquired animal botulism is mainly due to BoNT/C, D and the mosaic variants CD and DC, BoNT/CD being more prevalent in birds and BoNT/DC in cattle, whereas human botulism is more frequently in the types A, B and E, and to a lower extent, F. Botulism is not a contagious disease, since there is no direct transmission from diseased animals or man to a healthy subject. Botulism occurs via the environment, notably from food contaminated with spores and preserved in conditions favorable for growth and toxin production. The high prevalence of botulism types C, D and variants DC and CD in farmed and wild birds, and to a lower extent in cattle, raises the risk of transmission to human beings. However, human botulism is much rarer than animal botulism, and botulism types C and D are exceptional in humans. Only 15 cases or suspected cases of botulism type C and one outbreak of botulism type D have been reported in humans to date. In contrast, animal healthy carriers of group II, such as type E in fish of the northern hemisphere, and B4 in pigs, represent a more prevalent risk of botulism transmission to human subjects. Less common botulism types in animals but at risk of transmission to humans, can sporadically be observed, such as botulism type E in farmed chickens in France (1998-2002), botulism type B in cattle in The Netherlands (1977-1979), botulism types A and B in horses, or botulism type A in dairy cows (Egypt, 1976). In most cases, human and animal botulisms have distinct origins, and cross transmissions between animals and human beings are rather rare, accidental events. But, due to the severity of this disease, human and animal botulism requires a careful surveillance.
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http://dx.doi.org/10.3390/toxins12010017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7020394PMC
December 2019

Antibodies and Vaccines against Botulinum Toxins: Available Measures and Novel Approaches.

Toxins (Basel) 2019 09 12;11(9). Epub 2019 Sep 12.

Institut Pasteur, Département de Microbiologie, Unité des Toxines Bactériennes, 25 Rue du Docteur Roux, 75015 Paris, France.

Botulinum neurotoxin (BoNT) is produced by the anaerobic, Gram-positive bacterium . As one of the most poisonous toxins known and a potential bioterrosism agent, BoNT is characterized by a complex mode of action comprising: internalization, translocation and proteolytic cleavage of a substrate, which inhibits synaptic exocytotic transmitter release at neuro-muscular nerve endings leading to peripheral neuroparalysis of the skeletal and autonomic nervous systems. There are seven major serologically distinct toxinotypes (A-G) of BoNT which act on different substrates. Human botulism is generally caused by BoNT/A, B and E. Due to its extreme lethality and potential use as biological weapon, botulism remains a global public health concern. Vaccination against BoNT, although an effective strategy, remains undesirable due to the growing expectation around therapeutic use of BoNTs in various pathological conditions. This review focuses on the current approaches for botulism control by immunotherapy, highlighting the future challenges while the molecular underpinnings among subtypes variants and BoNT sequences found in non-clostridial species remain to be elucidated.
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http://dx.doi.org/10.3390/toxins11090528DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6783819PMC
September 2019

DABMA: A Derivative of ABMA with Improved Broad-Spectrum Inhibitory Activity of Toxins and Viruses.

ACS Med Chem Lett 2019 Aug 2;10(8):1140-1147. Epub 2019 Jul 2.

Service d'Ingénierie Moléculaire des Protéines (SIMOPRO), CEA, Université Paris-Saclay, LabEx LERMIT, 91191 Gif-sur-Yvette, France.

The small molecule ABMA has been previously shown to protect cells against multiple toxins and pathogens including virus, intracellular bacteria, and parasite. Its mechanism of action is directly associated with host endolysosomal pathway rather than targeting toxin or pathogen itself. However, the relationship of its broad-spectrum anti-infection activity and chemical structure is not yet resolved. Here, we synthesized a series of derivatives and compared their activities against diphtheria toxin (DT). Dimethyl-ABMA (DABMA), one of the most potent analogs with about 20-fold improvement in protection efficacy against DT, was identified with a similar mechanism of action to ABMA. Moreover, DABMA exhibited enhanced efficacy against toxin B (TcdB), lethal toxin (TcsL), Exotoxin A (PE) as well as Rabies and Ebola viruses. The results revealed a structure-activity relationship of ABMA, which is a starting point for its clinical development as broad-spectrum drug against existing and emerging infectious diseases.
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http://dx.doi.org/10.1021/acsmedchemlett.9b00155DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6691562PMC
August 2019

The pore structure of Clostridium perfringens epsilon toxin.

Nat Commun 2019 06 14;10(1):2641. Epub 2019 Jun 14.

College of Life and Environmental Sciences, University of Exeter, Stocker Road, Exeter, EX4 4QD, UK.

Epsilon toxin (Etx), a potent pore forming toxin (PFT) produced by Clostridium perfringens, is responsible for the pathogenesis of enterotoxaemia of ruminants and has been suggested to play a role in multiple sclerosis in humans. Etx is a member of the aerolysin family of β-PFTs (aβ-PFTs). While the Etx soluble monomer structure was solved in 2004, Etx pore structure has remained elusive due to the difficulty of isolating the pore complex. Here we show the cryo-electron microscopy structure of Etx pore assembled on the membrane of susceptible cells. The pore structure explains important mutant phenotypes and suggests that the double β-barrel, a common feature of the aβ-PFTs, may be an important structural element in driving efficient pore formation. These insights provide the framework for the development of novel therapeutics to prevent human and animal infections, and are relevant for nano-biotechnology applications.
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http://dx.doi.org/10.1038/s41467-019-10645-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6572795PMC
June 2019

Editorial: Pathogens and disease special issue ETOX18.

Pathog Dis 2019 03;77(2)

Bacterial Toxins, Institut Pasteur, Paris, France.

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http://dx.doi.org/10.1093/femspd/fty091DOI Listing
March 2019

Whole-Genome Sequences of a Cluster of 14 Unidentified Related sp. Strains from Human Clinical Samples and Type Strains of 3 Validated Species.

Microbiol Resour Announc 2019 Mar 21;8(12). Epub 2019 Mar 21.

HydroSciences Montpellier, CNRS, IRD, Université de Montpellier, CHU Nîmes, Nîmes, France

We report 17 draft genomes for 14 unidentified sp. strains closely related in 16S rRNA gene-based phylogeny and type strains of 3 species with the aims of deciphering relationships between related species, evaluating the accuracy of current thresholds for species delineation, and robustly describing new species in the genus.
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http://dx.doi.org/10.1128/MRA.01743-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6430328PMC
March 2019

Two Type E Isolates in France.

Toxins (Basel) 2019 03 1;11(3). Epub 2019 Mar 1.

Bacterial Toxins, Institut Pasteur, ERL 6002, 75015 Paris, France.

type E is a less frequently isolated type and has not previously been reported in France. We have characterized two recent type E isolates, 508.17 from the intestinal content of a calf that died of enterotoxemia, and 515.17 from the stool of a 60-year-old woman, subsequent to food poisoning, which contained the plasmid pCPPB-1 with variant iota toxin and enterotoxin genes.
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http://dx.doi.org/10.3390/toxins11030138DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6468631PMC
March 2019

Revisiting an old antibiotic: bacitracin neutralizes binary bacterial toxins and protects cells from intoxication.

FASEB J 2019 04 30;33(4):5755-5771. Epub 2019 Jan 30.

Institute of Pharmacology and Toxicology, University of Ulm Medical Center, Ulm, Germany.

The antibiotic bacitracin (Bac) inhibits cell wall synthesis of gram-positive bacteria. Here, we discovered a totally different activity of Bac: the neutralization of bacterial exotoxins. Bac prevented intoxication of mammalian cells with the binary enterotoxins Clostridium botulinum C2, C. perfringens ι, C. difficile transferase (CDT), and Bacillus anthracis lethal toxin. The transport (B) subunits of these toxins deliver their respective enzyme (A) subunits into cells. Following endocytosis, the B subunits form pores in membranes of endosomes, which mediate translocation of the A subunits into the cytosol. Bac inhibited formation of such B pores in lipid bilayers in vitro and in living cells, thereby preventing translocation of the A subunit into the cytosol. Bac preserved the epithelial integrity of toxin-treated CaCo-2 monolayers, a model for the human gut epithelium. In conclusion, Bac should be discussed as a therapeutic option against infections with medically relevant toxin-producing bacteria, including C. difficile and B. anthracis, because it inhibits bacterial growth and neutralizes the secreted toxins.-Schnell, L., Felix, I., Müller, B., Sadi, M., von Bank, F., Papatheodorou, P., Popoff, M. R., Aktories, K., Waltenberger, E., Benz, R., Weichbrodt, C., Fauler, M., Frick, M., Barth, H. Revisiting an old antibiotic: bacitracin neutralizes binary bacterial toxins and protects cells from intoxication.
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http://dx.doi.org/10.1096/fj.201802453RDOI Listing
April 2019

Why Are Botulinum Neurotoxin-Producing Bacteria So Diverse and Botulinum Neurotoxins So Toxic?

Toxins (Basel) 2019 01 11;11(1). Epub 2019 Jan 11.

Bacterial Toxins, Institut Pasteur, 75015 Paris, France.

Botulinum neurotoxins (BoNTs) are the most lethal toxins among all bacterial, animal, plant and chemical poisonous compounds. Although a great effort has been made to understand their mode of action, some questions are still open. Why, and for what benefit, have environmental bacteria that accidentally interact with their host engineered so diverse and so specific toxins targeting one of the most specialized physiological processes, the neuroexocytosis of higher organisms? The extreme potency of BoNT does not result from only one hyperactive step, but in contrast to other potent lethal toxins, from multi-step activity. The cumulative effects of the different steps, each having a limited effect, make BoNTs the most potent lethal toxins. This is a unique mode of evolution of a toxic compound, the high potency of which results from multiple steps driven by unknown selection pressure, targeting one of the most critical physiological process of higher organisms.
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http://dx.doi.org/10.3390/toxins11010034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6357194PMC
January 2019

Variability of Botulinum Toxins: Challenges and Opportunities for the Future.

Toxins (Basel) 2018 09 13;10(9). Epub 2018 Sep 13.

Institut Pasteur, Département de Microbiologie, Unité des Toxines Bactériennes, 25 Rue du Docteur Roux, 75015 Paris, France.

Botulinum neurotoxins (BoNTs) are the most potent known toxins, and are therefore classified as extremely harmful biological weapons. However, BoNTs are therapeutic drugs that are widely used and have an increasing number of applications. BoNTs show a high diversity and are divided into multiple types and subtypes. Better understanding of the activity at the molecular and clinical levels of the natural BoNT variants as well as the development of BoNT-based chimeric molecules opens the door to novel medical applications such as silencing the sensory neurons at targeted areas and dermal restoration. This short review is focused on BoNTs' variability and the opportunities or challenges posed for future clinical applications.
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http://dx.doi.org/10.3390/toxins10090374DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6162648PMC
September 2018

Enterotoxin: The Toxin Forms Highly Cation-Selective Channels in Lipid Bilayers.

Toxins (Basel) 2018 08 22;10(9). Epub 2018 Aug 22.

Bacterial Toxins, Institut Pasteur, 28 rue du Dr Roux, 75015 Paris, France.

One of the numerous toxins produced by is enterotoxin (CPE), a polypeptide with a molecular mass of 35.5 kDa exhibiting three different domains. Domain one is responsible for receptor binding, domain two is involved in hexamer formation and domain three has to do with channel formation in membranes. CPE is the major virulence factor of this bacterium and acts on the claudin-receptor containing tight junctions between epithelial cells resulting in various gastrointestinal diseases. The activity of CPE on Vero cells was demonstrated by the entry of propidium iodide (PI) in the cells. The entry of propidium iodide caused by CPE was well correlated with the loss of cell viability monitored by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test. CPE formed ion-permeable channels in artificial lipid bilayer membranes with a single-channel conductance of 620 pS in 1 M KCl. The single-channel conductance was not a linear function of the bulk aqueous salt concentration indicating that point-negative charges at the CPE channel controlled ion transport. This resulted in the high cation selectivity of the CPE channels, which suggested that anions are presumably not permeable through the CPE channels. The possible role of cation transport by CPE channels in disease caused by is discussed.
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http://dx.doi.org/10.3390/toxins10090341DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6162509PMC
August 2018

Expansion of the Clostridium perfringens toxin-based typing scheme.

Anaerobe 2018 Oct 20;53:5-10. Epub 2018 Apr 20.

Department of Pathology, Bacteriology and Avian Diseases, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium.

Clostridium perfringens causes many different histotoxic and enterotoxic diseases in humans and animals as a result of its ability to produce potent protein toxins, many of which are extracellular. The current scheme for the classification of isolates was finalized in the 1960s and is based on their ability to produce a combination of four typing toxins - α-toxin, β-toxin, ε-toxin and ι-toxin - to divide C. perfringens strains into toxinotypes A to E. However, this scheme is now outdated since it does not take into account the discovery of other toxins that have been shown to be required for specific C. perfringens-mediated diseases. We present a long overdue revision of this toxinotyping scheme. The principles for the expansion of the typing system are described, as is a mechanism by which new toxinotypes can be proposed and subsequently approved. Based on these criteria two new toxinotypes have been established. C. perfringens type F consists of isolates that produce C. perfringens enterotoxin (CPE), but not β-toxin, ε-toxin or ι-toxin. Type F strains will include strains responsible for C. perfringens-mediated human food poisoning and antibiotic associated diarrhea. C. perfringens type G comprises isolates that produce NetB toxin and thereby cause necrotic enteritis in chickens. There are at least two candidates for future C. perfringens toxinotypes, but further experimental work is required before these toxinotypes can formally be proposed and accepted.
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http://dx.doi.org/10.1016/j.anaerobe.2018.04.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6195859PMC
October 2018

Evidence of Clostridium perfringens epsilon toxin associated with multiple sclerosis.

Mult Scler 2019 04 21;25(5):653-660. Epub 2018 Apr 21.

College of Life and Environmental Sciences, University of Exeter, Exeter, UK.

Background: It was recently reported that, using Western blotting, some multiple sclerosis (MS) patients in the United States had antibodies against epsilon toxin (Etx) from Clostridium perfringens, suggesting that the toxin may play a role in the disease.

Objective: We investigated for serum antibodies against Etx in UK patients with clinically definite multiple sclerosis (CDMS) or presenting with clinically isolated syndrome (CIS) or optic neuritis (ON) and in age- and gender-matched controls.

Methods: We tested sera from CDMS, CIS or ON patients or controls by Western blotting. We also tested CDMS sera for reactivity with linear overlapping peptides spanning the amino acid sequence (Pepscan) of Etx.

Results: Using Western blotting, 24% of sera in the combined CDMS, CIS and ON groups ( n = 125) reacted with Etx. In the control group ( n = 125), 10% of the samples reacted. Using Pepscan, 33% of sera tested reacted with at least one peptide, whereas in the control group only 16% of sera reacted. Out of 61 samples, 21 (43%) were positive to one or other testing methodology. Three samples were positive by Western blotting and Pepscan.

Conclusion: Our results broadly support the previous findings and the role of Etx in the aetiology of MS warrants further investigation.
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http://dx.doi.org/10.1177/1352458518767327DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6439943PMC
April 2019

Human alpha-defensin-1 protects cells from intoxication with Clostridium perfringens iota toxin.

Pathog Dis 2018 03;76(2)

Institute of Pharmacology and Toxicology, University of Ulm Medical Center, 89081 Ulm, Germany.

Iota toxin is produced by Clostridium perfringens type E strains and associated with diarrhea in cattle and lambs. This binary protein toxin comprises the enzyme component iota a (Ia), which ADP-ribosylates G-actin, and the separate transport component iota b (Ib), which delivers Ia into the cytosol of target cells. Ib binds to cell receptors and forms biologically active toxin complexes with Ia, which cause rounding of adherent cells due to the destruction of the actin cytoskeleton. Here, we report that the human peptide α-defensin-1 protects cultured cells including human colon cells from intoxication with iota toxin. In contrast, the related ß-defensin-1 had no effect, indicating a specific mode of action. The α-defensin-1 did not inhibit ADP-ribosylation of actin by Ia in vitro. Pretreatment of Ib with α-defensin-1 prior to addition of Ia prevented intoxication. Additionally, α-defensin-1 protected cells from cytotoxic effects mediated by Ib in the absence of Ia, implicating that α-defensin-1 interacts with Ib to prevent the formation of biologically active iota toxin on cells. In conclusion, the findings contribute to a better understanding of the functions of α-defensin-1 and suggest that this human peptide might be an attractive starting point to develop novel pharmacological options to treat/prevent diseases associated with iota toxin-producing Clostridium perfringens strains.
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http://dx.doi.org/10.1093/femspd/fty022DOI Listing
March 2018

Botulinum Neurotoxins: Still a Privilege of Clostridia?

Authors:
Michel R Popoff

Cell Host Microbe 2018 02;23(2):145-146

Bacterial Toxins, Institut Pasteur, Paris, France. Electronic address:

Botulinum neurotoxins (BoNTs) are potent bacterial toxins mostly produced by genetically diverse clostridial strains. Recently, BoNT variants have been reported in non-clostridial strains. In this issue of Cell Host & Microbe, Zhang et al. (2018) describe a novel BoNT in Entecoccus faecium.
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http://dx.doi.org/10.1016/j.chom.2018.01.014DOI Listing
February 2018

"Bacterial Toxins" Section in the Journal Toxins: A Fantastic Multidisciplinary Interplay between Bacterial Pathogenicity Mechanisms, Physiological Processes, Genomic Evolution, and Subsequent Development of Identification Methods, Efficient Treatment, and Prevention of Toxigenic Bacteria.

Authors:
Michel R Popoff

Toxins (Basel) 2018 01 18;10(1). Epub 2018 Jan 18.

Institut Pasteur, Unité des Bactéries Anaérobies et Toxines, 25 Avenue du Docteur Roux, 75015 Paris, France.

Toxins are powerful pathogenicity factors produced by certain bacteria, fungi, animals, and plants which mediate drastic interactions of these pathogens on the organism host[...].
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http://dx.doi.org/10.3390/toxins10010044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5793131PMC
January 2018