Clostridium Botulinum Research Articles

Exploring the therapeutic potential of cinnamon and clove essential oils for mitigating foodborne diseases: bioactivity and molecular interactions

Foodborne diseases remain a global concern due to their widespread impact on public health. Contaminated food can lead to outbreaks of illness that affect millions of people each year, leading to significant economic burdens and strain on healthcare systems. This study delves into the potential of cinnamon and clove essential oils to combat foodborne diseases. Essential oils extracted via the Soxhlet method using ethanol as solvent, these oils exhibit potent antibacterial, anti-inflammatory, antioxidant and anticancer properties. Gas chromatography-mass spectrometry analysis was performed to analyze the chemical constituents in both essential oils. Employing the Lipinski rule, bioactive compounds are screened and docked against a viral replication responsible protein of norovirus and the neurotoxin (botulinum toxin) of Clostridium botulinum both responsible for severe foodborne illnesses. Acetyleugenol demonstrates strong binding affinity, while the pyrimidine derivative from cinnamon sourced from Mbeya, Tanzania forms pi-sulfur interactions with both target molecules. This research emphasizes how essential oils confront foodborne diseases, granting valuable perspectives on potential compounds for extended examination and therapeutic interventions.

Comprehensive identification of pathogenic microbes and antimicrobial resistance genes in food products using nanopore sequencing-based metagenomics

Foodborne pathogens, particularly antimicrobial-resistant (AMR) bacteria, remain a significant threat to global health. Given the limitations of conventional culture-based approaches, which are limited in scope and time-consuming, metagenomic sequencing of food products emerges as a promising solution. This method provides a fast and comprehensive way to detect the presence of pathogenic microbes and antimicrobial resistance genes (ARGs). Notably, nanopore long-read sequencing provides more accurate bacterial taxonomic classification in comparison to short-read sequencing. Here, we revealed the impact of food types and attributes (origin, retail place, and food processing methods) on microbial communities and the AMR profile using nanopore metagenomic sequencing. We analyzed a total of 260 food products, including raw meat, sashimi, and ready-to-eat (RTE) vegetables. Clostridium botulinum, Acinetobacter baumannii, and Vibrio parahaemolyticus were identified as the top three foodborne pathogens in raw meat and sashimi. Importantly, even with low pathogen abundance, higher percentages of samples containing carbapenem and cephalosporin resistance genes were identified in chicken and RTE vegetables, respectively. In parallel, our results demonstrated that fresh, peeled, and minced foods exhibited higher levels of pathogenic bacteria. In conclusion, this comprehensive study offers invaluable data that can contribute to food safety assessments and serve as a basis for quality indicators.

Botulism in fish: a review.

Published information about fish botulism is scant. We review here the current literature on fish botulism. Freshwater fish are susceptible to botulism. Only anecdotal evidence exists about possible botulism cases in saltwater fish. With only a few exceptions, the etiology of all cases of fish botulism reported is Clostridium botulinum type E, although fish are sensitive to, and may carry, various C. botulinum types. Clinical signs of botulism in fish include loss of equilibrium and motion, abducted opercula, open mouths, dark pigmentation, and head up/tail down orientation in which attempts to swim result in breaching the surface of the water. Dark pigmentation is thought to be associated with acetylcholine imbalance in botulinum neurotoxin (BoNT)-affected fish. Rarely, but similar to the situation in other animal species, fish can recover from botulism. Fish botulism can cause secondary outbreaks of the disease in birds, as botulism-affected fish stand out from normal fish, and are selectively preyed upon by fish-eating birds, which thus become intoxicated by the BoNT present in sick fish. The source of BoNT in fish has not been definitively confirmed. Fish may ingest C. botulinum spores that then germinate in their digestive tract, but the possibility that fish ingest preformed BoNT from the environment (e.g., dead fish, shellfish, insects) cannot be ruled out. The presumptive diagnosis of botulism in fish is established based on clinical signs, and as in other species, confirmation should be based on detection of BoNT in intestinal content, liver, and/or serum of affected fish.

Efficacy and Safety Analysis of Botox Application and Iatrogenic Botulism: Panacea or Peril?

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.

Emulsified and Nanoemulsified Essential Oils in the Control of Clostridium botulinum and Clostridium sporogenes in Mortadella.

Clostridium botulinum is a foodborne pathogen responsible for severe neuroparalytic disease associated with the ingestion of pre-formed toxin in food, with processed meats and canned foods being the most affected. Control of this pathogen in meat products is carried out using the preservative sodium nitrite (NaNO2), which in food, under certain conditions, such as thermal processing and storage, can form carcinogenic compounds. Therefore, the objective was to use nanoemulsified essential oils (EOs) as natural antimicrobial agents, with the aim of reducing the dose of NaNO2 applied in mortadella. The antimicrobial activity of nanoemulsions prepared with mixtures of EOs of garlic, clove, pink pepper, and black pepper was evaluated on endospores and vegetative cells of C. botulinum and Clostridium sporogenes (surrogate model) inoculated in mortadella prepared with 50 parts per million NaNO2. The effects on the technological (pH, water activity, and color) and sensory characteristics of the product were also evaluated. The combinations of EOs and their nanoemulsions showed sporicidal effects on the endospores of both tested microorganisms, with no counts observed from the 10th day of analysis. Furthermore, bacteriostatic effects on the studied microorganisms were observed. Regarding the technological and sensorial characteristics of the product, the addition of the combined EOs had a negative impact on the color of the mortadella and on the flavor/aroma. Despite the strong commercial appeal of adding natural preservatives to foods, the effects on flavor and color must be considered. Given the importance of controlling C. botulinum in this type of product, as well as the reduction in the amount of NaNO2 used, this combination of EOs represents a promising antimicrobial alternative to this preservative, encouraging further research in this direction.

Evaluation of long-term immune response in cattle to botulism using a recombinant E. coli bacterin formulated with Montanide™ ISA 50 and aluminum hydroxide adjuvants

Botulism is a severe disease caused by potent botulinum neurotoxins (BoNTs) produced by Clostridium botulinum. This disease is associated with high-lethality outbreaks in cattle, which have been linked to the ingestion of preformed BoNT serotypes C and D, emphasizing the need for effective vaccines. The potency of current commercial toxoids (formaldehyde-inactivated BoNTs) is assured through tests in guinea pigs according to government regulatory guidelines, but their short-term immunity raises concerns. Recombinant vaccines containing the receptor-binding domain have demonstrated potential for eliciting robust protective immunity. Previous studies have demonstrated the safety and effectiveness of recombinant E. coli bacterin, eliciting high titers of neutralizing antibodies against C. botulinum and C. perfringens in target animal species. In this study, neutralizing antibody titers in cattle and the long-term immune response against BoNT/C and D were used to assess the efficacy of the oil-based adjuvant compared with that of the aluminum hydroxide adjuvant in cattle. The vaccine formulation containing Montanide™ ISA 50 yielded significantly higher titers of neutralizing antibody against BoNT/C and D (8.64 IU/mL and 9.6 IU/mL, respectively) and induced an immune response that lasted longer than the response induced by aluminum, extending between 30 and 60 days. This approach represents a straightforward, cost-effective strategy for recombinant E. coli bacterin, enhancing both the magnitude and duration of the immune response to botulism.

Microbiological risk assessment and resistome analysis from shotgun metagenomics of bovine colostrum microbiome

Colostrum is known for its nutraceutical qualities, probiotic attributes, and health benefits. The aim of this study was to profile colostrum microbiome from bovine in rural sites of a developing country. The focus was on microbiological safety assessments and antimicrobial resistance, taking into account the risks linked with the consumption of raw colostrum. Shotgun sequencing was employed to analyze microbiome in raw buffalo and cow colostrum. Alpha and beta diversity analyses revealed increased inter and intra-variability within colostrum samples' microbiome from both livestock species. The colostrum microbiome was mainly comprised of bacteria, with over 90% abundance, whereas fungi and viruses were found in minor abundance. Known probiotic species, such as Leuconostoc mesenteroides, Lactococcus lactis, Streptococcus thermophilus, and Lactobacillus paracasei, were found in the colostrum samples. A relatively higher number of pathogenic and opportunistic pathogenic bacteria were identified in colostrum from both animals, including clinically significant bacteria like Clostridium botulinum, Pseudomonas aeruginosa, Escherichia coli, and Listeria monocytogenes. Binning retrieved 11 high-quality metagenome-assembled genomes (MAGs), with three MAGs potentially representing novel species from the genera Psychrobacter and Pantoea. Notably, 175 antimicrobial resistance genes (ARGs) and variants were detected, with 55 of them common to both buffalo and cow colostrum metagenomes. These ARGs confer resistance against aminoglycoside, fluoroquinolone, tetracycline, sulfonamide, and peptide antibiotics. In conclusion, this study describes a thorough overview of microbial communities in buffalo and cow colostrum samples. It emphasizes the importance of hygienic processing and pasteurization in minimizing the potential transmission of harmful microorganisms linked to the consumption of colostrum.

Botulinum neurotoxin X lacks potency in mice and in human neurons.

The family of botulinum neurotoxins comprises the most potent toxins known to humankind. New members of this family of protein toxins as well as more distantly related homologs are being identified. The discovery of BoNT/X via bioinformatic screen in 2017 as a putative new BoNT serotype raised concern about its potential as a pathogenic agent with no available countermeasures. This study for the first time assessed both recombinantly produced and native purified BoNT/X for its vertebrate neurotoxicity.

From Toxin to Treatment: A Narrative Review on the Use of Botulinum Toxin for Autonomic Dysfunction.

Since its regulatory approval over a half-century ago, botulinum toxin has evolved from one of the most potent neurotoxins known to becoming routinely adopted in clinical practice. Botulinum toxin, a highly potent neurotoxin produced by Clostridium botulinum, can cause botulism illness, characterized by widespread muscle weakness due to inhibition of acetylcholine transmission at neuromuscular junctions. The observation of botulinum toxin's anticholinergic properties led to the investigation of its potential benefits for conditions with an underlying etiology of cholinergic transmission, including autonomic nervous system dysfunction. These conditions range from disorders of the integument to gastrointestinal and urinary systems. Several formulations of botulinum toxin have been developed and tested over time, significantly increasing the availability of this treatment for appropriate clinical use. Despite the accelerated and expanded use of botulinum toxin, there lacks an updated comprehensive review on its therapeutic use, particularly to treat autonomic dysfunction. This narrative review provides an overview of the effect of botulinum toxin in the treatment of autonomic dysfunction and summarizes the different formulations and dosages most widely studied, while highlighting reported outcomes and the occurrence of any adverse events.

Equine grass sickness is associated with major abnormalities in the ultrastructure of skeletal neuromuscular junctions.

Equine grass sickness (EGS) is a frequently fatal multisystem neuropathy of equids. The aetiology is unknown; proposed causes include toxicoinfection with Clostridium botulinum and a mycotoxicosis. The effect of EGS on the organisation and structural integrity of the skeletal neuromuscular junction (NMJ), the target of botulinum neurotoxins (BoNTs), is unknown. To compare the organisation and structural integrity of skeletal NMJs from EGS horses, control horses and one horse with a presumptive diagnosis of botulism. Blinded, retrospective case control. NMJs in samples of diaphragm or intercostal muscle from six EGS horses, three control horses and one equine botulism case were compared using electron microscopy, morphometry and confocal light microscopy. A significantly higher percentage of EGS NMJs had abnormal morphology (EGS 72.2%, 95% CI 55.6-84.4; Controls 6.9%, 1.7-23.8; OR 35.1, 8.47-244.8; p < 0.001). EGS NMJs had a significantly lower mean volume fraction occupied by synaptic vesicles (SVs) (EGS 18.7%, 12.6-28.0; Controls 36.3%, 20.8-63.4; p = 0.024). EGS NMJs had evidence of accelerated SV exocytosis and SV depletion, accumulation of neurofilament-like material in terminal boutons and/or bouton degeneration. NMJs from the botulism horse had dense packing of SVs towards the presynaptic membrane active zone, consistent with BoNT intoxication, but had absence of the abnormalities identified in EGS NMJs. Group sizes were limited by difficulties obtaining suitably processed samples. Ages of control and EGS horses differed. Botulism was diagnosed based on clinical and post mortem findings. EGS is associated with major changes in skeletal NMJ ultrastructure that are inconsistent with the effects of BoNTs. SV depletion may reflect increased exocytosis coupled with reduced repopulation of SVs via anterograde axonal transport and endocytosis, consistent with the action of an excitatory presynaptic toxin and/or neurotransmitter reuptake inhibitor. Skeletal NMJs represent a previously unrecognised target for the toxin that causes EGS.

Interaction of Fe2O3 and Fe3O4 Nanoparticle with Pathogenic Bacteria: A In-silico Molecular Mechanism Study

Background:: Magnetic materials like iron, nickel, and cobalt have been a subject of interest among the scientific and research community for centuries. Owing to their unique properties, they are prevalent in the mechanical and electronic industries. In recent times, magnetic materials have undeniable applications in biotechnology and nanomedicine. Bacteria like Salmonella enterica, Clostridium botulinum, Bacillus subtilis, etc, pose a hazard to human health and livestock. This ultimately leads to huge yields and economic losses on a global scale. Antimicrobial resistance has become a significant public health concern in recent years, with the increasing prevalence of drugresistant infections posing a significant threat to global health. Many coherent studies have successfully reported magnetic metal oxide nanoparticles to be highly selective, specific, and effective in neutralizing pathogens through various mechanisms like cell membrane disruption, direct contact-mediated killing, or by generating Reactive Oxygen Species (ROS) and numerous costimulatory and inflammatory cytokines. Therefore, we explored the inhibitory effects of iron oxide nanoparticles (NPs) on various pathogenic bacteria via an in-silico approach. This method helped us to understand the active sites where the iron oxide NPs bind with the bacterial proteins. Methods:: The 3D crystal structures of all the pathogenic proteins of Streptococcus pneumoniae, Pseudomonas aeruginosa, Vibrio cholerae, Salmonella enterica, Shigella flexneri, Clostridium botulinum and nanoparticles (Fe2O3 and Fe3O4) under study were downloaded from RCSB PDB and ChemSpider official websites respectively. It was followed by the in-silico molecular Docking using PyRx and AutoDock Vina and analyzed on LigPlot. Results:: This study interprets the efficacy of the Fe2O3 and Fe3O4 nanoparticles against all the test bacteria. At the same time, Fe2O3 and Fe3O4 formed the most stable complexes with cholera enterotoxin subunit B and lectin II (PA-IIL) mutant S23A of Pseudomonas aeruginosa, respectively. Conclusion:: As in this era of AMR, researchers have been exploring alternative strategies to combat bacterial infections, including using magnetic nanoparticles as a potential treatment. They possess unique physical and chemical properties that make them attractive candidates for antimicrobial therapy, including their ability to penetrate bacterial biofilms and selectively target pathogenic bacteria while leaving healthy cells unharmed. This study examined the inhibitory effects of iron oxide (magnetic) nanoparticles, namely Fe2O3 and Fe3O4, on various bacterial proteins involved in cell-to-cell interactions and pathogenesis.

Pathogenic Escherichia coli (E. coli) food borne outbreak: Detection methods and controlling measures

Food-borne illnesses and diseases are major threats to human health. Common food borne pathogens include bacteria such as Escherichia coli Serogroup (O157), Bacillus cereus, Clostridium botulinum, Campylobacter sp., C. perfringens, some Listeria monocytogenes, Salmonella spp., Shigella spp., Staphylococcus aureus, and Vibrio spp. etc. Pathogenic E. coli infection usually causes severe diarrhea. Escherichia coli is the most common cause of acute urinary tract infections as well as urinary tract sepsis. Diarrheagenic E. coli pathotypes can be passed in the feces of humans and other animals. Safeguarding public health during mass gatherings is a big challenge. Despite the recent advances in food preservation techniques and food safety, significant disease outbreaks linked to food borne pathogens such as bacteria, fungi, and viruses still occur worldwide indicating that these pathogens still constitute significant risks to public health. However, at present the conventional method for E. coli detection requires several days, especially in cases where E. coli concentrations are low. Enrichment is a commonly used method for bacterial isolation to increase the cell counts of target bacteria. However, traditional culturing method is also a slow and laborious process requiring a series of steps and may require the use of adjunct methods (e.g. biochemical, serological, nucleic acid-based methods for conclusive identification) and can take up to a week for bacteria. The culture-based approaches coupled with other methods such as PCR, immunoassays, bacteriophages, NGS, Biosensors, and MALDI TOF MS are increasingly being used for the detection and identification of food borne pathogens. Nucleic acid-based methods such as Real-Time PCR and vPCR combined with sequencing approaches are more widely used than immunoassay and NGS-based approaches for pathogen detection. Nanobiotechnology is the latest approach for the detection of pathogens. Bacteriophages are one such unique biological entity that showed excellent host selectivity and have been actively used as recognition probes for pathogen detection. Antibiotics are efficient, powerful, and the most commonly used treatment against pathogenic E. coli. Other ways to treat or prevent E. coli contamination, such as probiotics, antimicrobial nanoparticles, and radiation treatment had been reported.

Simultaneous detection of multiple spoilage bacteria using a fluorogenic loop-mediated isothermal amplification-based multi-sample microfluidic chip

Spoilage bacteria, which ubiquitously contribute to food spoilage, pose a global public health risk. In this study, we developed a centrifugal microfluidic chip integrating fluorogenic loop-mediated isothermal amplification (LAMP) to detect six important spoilage bacteria: Bacillus cereus, Clostridium botulinum, Photobacterium phosphoreum, Pseudomonas putida, Streptococcus agalactiae, and Shewanella putrefaciens. The LAMP reaction components were pre-mixed (primers pre-embedded in each reaction chamber of the chip) and evenly pipetted into the two inlet holes, enabling the simultaneous analysis of six genetic targets from a single sample. This facilitates the automated detection of up to four samples simultaneously using a single chip. The LAMP reactions were conducted in a 5-μL volume at 65 °C for 60 min, achieving limits of detection ranging from 10−1 to 10−2 pg/μL of bacterial genomic DNA. Almost all coefficients of variation for time-to-positive values consistently remained under 10% when genomic DNA was used at concentrations within the limits of detection for the six bacteria. This demonstrated excellent reproducibility across replicates. The clinical sensitivity and specificity for field sample detection were 93.3% and 97.5%, respectively. This method offers ease of operation, rapid analysis, and minimal reagent usage and has great potential for the on-site detection of multiple spoilage bacteria.

Commercial farmed swine harbour a variety of pathogenic bacteria and antimicrobial resistance genes.

Introduction. The northern region of Thailand serves as a crucial area for swine production, contributing to the Thai community food supply. Previous studies have highlighted the presence of foodborne bacterial pathogens originating from swine farms in this region, posing a threat to both human and animal health.Gap statement. Multiple swine bacterial pathogens have been studied at a species level, but the distribution and co-occurrence of bacterial pathogens in agricultural swine has not been well established.Aim. Our study employed the intestinal scraping technique to directly examine the bacterial micro-organisms interacting with the swine host.Methodology. We used shotgun metagenomic sequencing to analyse the bacterial pathogens inhabiting the caecal microbiome of swine from five commercial farms in northern Thailand.Results. A variety of pathogenic and opportunistic bacteria were identified, including Escherichia coli, Clostridium botulinum, Staphylococcus aureus and the Corynebacterium genus. From a One Health perspective, these species are important foodborne and opportunistic pathogens in both humans and agricultural animals, making swine a critical pathogen reservoir that can cause illness in humans, especially farm workers. Additionally, the swine caecal microbiome contains commensal bacteria such as Bifidobacterium, Lactobacillus and Faecalibacterium, which are associated with normal physiology and feed utilization in healthy swine. Antimicrobial resistance genes were also detected in all samples, specifically conferring resistance to tetracycline and aminoglycosides, which have historically been used extensively in swine farming.Conclusion. The findings further support the need for improved sanitation standards in swine farms, and additional monitoring of agricultural animals and farm workers to reduce contamination and improved produce safety for human consumption.

The Latent Threat in Wild Birds: Clostridium botulinum.

Avian botulism caused by Clostridium botulinum emerged in 1910, affecting birds across North America, leading to severe outbreaks exacerbated by climate change, decreasing water levels, and inadequate wastewater management. While deadly for birds, its epidemiological impact on humans and other animals remains limited. Despite its significance, understanding and controlling the disease remain challenging. This review delves into the pathogen's epidemiology in wild bird populations, exploring the transmission, pathogenicity, clinical symptoms, diagnosis and treatment. The disease's growing concern in wild birds relates to the bacterium's adaptability and expansive spread, evident through genetic similarities among strains across countries. Outbreaks are influenced by environmental factors such as temperature and soil characteristics. Wild birds inadvertently transmit the bacterium, perpetuating the cycle through carcasses and flies. Some species suffer severely, while others, like scavengers, show resistance. Understanding disease mechanisms, involving potential toxin ingestion or internal production, remains ongoing. Clinical signs vary, affecting diverse bird orders. Diagnostic methods evolve, with treatment success varying among affected populations. Prevention and surveillance take precedence due to treatment challenges, emphasising population-based strategies and preventive measures to manage the widespread presence of C. botulinum.

Botulism is a Rare but Serious Disease

Botulism is a rare but serious disease caused by the anaerobic bacterium Clostridium botulinum, which produces spores and a toxin. It is very widespread in nature, in the ground and in the digestion of mice, rats and cats. The toxin is often created in rotten food, and the most common sources of infection for humans are canned goods and dried meat products. It is also possible to get the disease if an injured spot on the skin comes into contact with the causative agent of the disease. The disease is not contagious, but occurs suddenly and attacks the nerves.

Botulism in Two Juvenile Wild Red Foxes (Vulpes vulpes) in Somerset, England, UK.

Two juvenile red foxes (Vulpes vulpes) were euthanased because of severe nervous signs and paralysis. Detailed postmortem examinations were carried out with bacteriology, histology, and Clostridium botulinum toxin screening, which confirmed botulism as the cause of the clinical signs.

Closed genome sequence of Clostridium botulinum type B1 strain isolated from an infant botulism case in the United States.

We present the closed genome sequence of the Clostridium botulinum BT-22100019 strain isolated from the stool specimen of an infant diagnosed with botulism. With 4.33-Mb genome size and 28.0% G + C content, the bont/B1 gene encoded for botulinum neurotoxin serotype B was found on a 262 kb plasmid arranged in a ha+ orfx - cluster.

Complete genomes of Clostridium botulinum type B(F) isolates associated with a 1995 foodborne botulism outbreak from commercial pâté reveals a recombination event disrupting the ntnh gene.

Foodborne botulism is a neuroparalytic disease caused by ingestion of foods contaminated with botulinum neurotoxin (BoNT), produced by Clostridium botulinum. In 1995 a husband and wife from Québec, Canada, were hospitalized for several months with prolonged muscle paralysis after ingesting a commercial pâté de campagne. Examination of faecal samples from both patients and the pâté produced viable Group I (proteolytic) C. botulinum type B from each of the three samples. Whole genome sequencing revealed that all three isolates contain identical bont/B5 and bont/F2 genes encoded on a plasmid. Both faecal isolate genomes were identical in chromosome and plasmid length, as well as gene content. The genome of the pâté isolate was nearly identical to that of the faecal isolates with the notable difference of a missing 13-gene insertion on the bont/B5 cluster disrupting the ntnh gene. Examination of the insertion revealed several mobile genetic elements that participate in recombination.

Investigation of a Clostridium botulinum mass poisoning and lessons learned, Ivory Coast, 2023

Investigation of a Clostridium botulinum mass poisoning and lessons learned, Ivory Coast, 2023