RESUMEN
Cooked, uncured meat products packaged under reduced oxygen packaging conditions require the control of anaerobic and facultative anaerobic pathogens if they are held at temperatures greater than 3°C at retail or consumer level. The objective of this study was to determine the inhibition of Listeria monocytogenes and Clostridium botulinum in cooked, uncured shredded turkey and pork formulated with synthetic or clean-label antimicrobials. Treatments of shredded meat products were prepared with or without antimicrobials using turkey thigh or breast that were cooked to 85°C, shredded, and chilled before inoculation with the target pathogen. L. monocytogenes inoculated samples were stored at 7.2°C, whereas C. botulinum samples were stored at 12.8°C; triplicate samples were assayed every 2 weeks. In the first set of experiments, L. monocytogenes populations increased 2 to 3 logs within 2 weeks of storage at 7.2°C in both meat control treatments without antimicrobials and in pork with 4% lactate-diacetate blend (LD). A 1-log increase was observed in turkey with 4% LD and Pork with 2% cultured dextrose-vinegar-rosemary (CDVR) under the same storage conditions; a 1-log increase was observed in turkey with CDVR at 4 weeks. The second set of experiments tested the effect of pH reduction (to less than 5.5 by the addition of 0.5% citric acid) in combination with 2% CDVR when added to the brine precook or postcook during shredding. Populations of L. monocytogenes increased 4-log within 2 and 4 weeks at 7.2°C for the control turkey and pork formulations, respectively. No growth was observed in 12 weeks for any antimicrobial CDVR-CA treatments regardless of how antimicrobial was added. Similarly, botulinum toxin was detected in both control treatments at week 2 at 12.8°C, but no toxicity was observed in either antimicrobial treatment through 12 weeks. These data suggest that a combination of 2% cultured dextrose-vinegar-rosemary extract plus 0.5% citric acid to reduce pH inhibits the growth of L. monocytogenes and toxin production of C. botulinum in uncured shredded turkey and pork products stored under mild temperature abuse conditions for up to 12 weeks in reduced oxygen packaging.
Asunto(s)
Clostridium botulinum , Recuento de Colonia Microbiana , Microbiología de Alimentos , Listeria monocytogenes , Oxígeno , Pavos , Listeria monocytogenes/efectos de los fármacos , Animales , Clostridium botulinum/efectos de los fármacos , Porcinos , Humanos , Embalaje de Alimentos/métodos , Productos de la Carne/microbiología , Conservación de Alimentos/métodos , Contaminación de Alimentos/análisis , TemperaturaRESUMEN
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.
Asunto(s)
Clostridium botulinum , Clostridium , Productos de la Carne , Aceites Volátiles , Aceites Volátiles/farmacología , Clostridium botulinum/efectos de los fármacos , Productos de la Carne/microbiología , Clostridium/efectos de los fármacos , Microbiología de Alimentos , Nitrito de Sodio/farmacología , Emulsiones , Humanos , Conservación de Alimentos/métodos , Esporas Bacterianas/efectos de los fármacos , Conservantes de Alimentos/farmacología , Gusto , Antibacterianos/farmacologíaRESUMEN
Clostridium botulinum produces botulinum neurotoxin (BoNT), which is the most toxic known protein and the causative agent of human botulism. BoNTs have similar structures and functions, comprising three functional domains: catalytic domain (L), translocation domain (HN), and receptor-binding domain (Hc). In the present study, BoNT/E was selected as a model toxin to further explore the immunological significance of each domain. The EL-HN fragment (L and HN domains of BoNT/E) retained the enzymatic activity without in vivo neurotoxicity. Extensive investigations showed EL-HN functional fragment had the highest protective efficacy and contained some functional neutralizing epitopes. Further experiments demonstrated the EL-HN provided a superior protective effect compared with the EHc or EHc and EL-HN combination. Thus, the EL-HN played an important role in immune protection against BoNT/E and could provide an excellent platform for the design of botulinum vaccines and neutralizing antibodies. The EL-HN has the potential to replace EHc or toxoid as the optimal immunogen for the botulinum vaccine.
Asunto(s)
Anticuerpos Neutralizantes/inmunología , Vacunas Bacterianas/administración & dosificación , Vacunas Bacterianas/inmunología , Botulismo/inmunología , Botulismo/prevención & control , Clostridium botulinum/inmunología , Neurotoxinas/toxicidad , Animales , Clostridium botulinum/efectos de los fármacos , Modelos Animales de Enfermedad , Femenino , Humanos , Ratones , Sustancias Protectoras/administración & dosificación , SerogrupoRESUMEN
Clostridium sporogenes has been widely used as a surrogate for proteolytic C. botulinum for validating thermal processes in low-acid cans. To limit the intensity of heat treatments, industrials must use other ways of control as an association of acidic and saline environment after a low heat treatment. The probability of growth of pH (7-4.4), sodium chloride concentration (0-11%) and heat treatment (80°C-10 min; 100°C-1.5 min and 5.2 min) were studied on C. sporogenes PA 3679 spores and vegetative cells. Vegetative cells or heat-treated spores were inoculated in PYGm broth at 30 °C for 48 days in anaerobic conditions. Vegetative cells growth (pH 4.6-pH 4.5; 7%-8% NaCl) range is larger than the spore one (pH 5.2-pH 5.0; 6%-7% NaCl). Spores germination and outgrowth rage is decreased if the spores are heat-treated at 100 °C for 1.5 min (pH 5.5-5.3; 4%-5% NaCl) and 5.2 min (pH 5.7-5.3; 4%-5% NaCl). The C. sporogenes PA 3679 spores germination and outgrowth is impacted by their physiological state. The synergic interaction between environmental factors (pH and NaCl) and the physiological state (vegetative cells and spores) opening new possibilities for optimizing food formulation processes to manage the risks of C. sporogenes spoilage.
Asunto(s)
Clostridium/crecimiento & desarrollo , Conservación de Alimentos/métodos , Alimentos en Conserva/microbiología , Cloruro de Sodio/farmacología , Esporas Bacterianas/crecimiento & desarrollo , Clostridium/efectos de los fármacos , Clostridium botulinum/efectos de los fármacos , Clostridium botulinum/crecimiento & desarrollo , Contaminación de Alimentos/análisis , Microbiología de Alimentos , Conservación de Alimentos/instrumentación , Calor , Concentración de Iones de Hidrógeno , Viabilidad Microbiana , Cloruro de Sodio/análisisRESUMEN
Botulinum neurotoxins (BoNTs) are the most toxic substances known to humankind and are the causative agents of the neuroparalytic disease botulism. Despite the overall importance of BoNTs in public health and safety, as a bioterrorism concern, and in pharmaceutical development, little is known about the molecular mechanisms mediating BoNT stability and degradation in various environments. Previous studies using Clostridium botulinum strain ATCC 3502 revealed that high levels of arginine (20 g/liter) repressed BoNT production approximately 1,000-fold. In the present study, the mechanisms of toxin reduction in arginine-enriched cultures of C. botulinum strain Hall A-hyper, which we have previously genetically manipulated using ClosTron technology, were explored. Cultures were grown in toxin production medium (TPM) and TPM enriched with arginine. Cultures were analyzed for growth (optical density at 600 nm [OD600]), changes in pH, and BoNT formation and stability. Our data indicate that arginine enrichment of C. botulinum strain Hall A-hyper cultures results in a pH shift that induces pH-dependent posttranslational control mechanisms. We further show that independent of arginine, maintenance of an acidic culture pH during growth of C. botulinum strain Hall A-hyper plays a central role in toxin stability and that an extracellular metalloprotease produced by the culture results in BoNT degradation at pH levels between â6.5 and 8.0. IMPORTANCE Botulinum neurotoxin (BoNT) is a public health and bioterrorism concern as well as an important and widely used pharmaceutical, yet the regulation of its synthesis by BoNT-producing clostridia is not well understood. This paper highlights the role of environmentally controlled posttranslational regulatory mechanisms influencing processing and stability of biologically active BoNTs produced by C. botulinum. The results of this work will help enhance public health and safety measures and our ability to evaluate safety risks of novel BoNTs and improve production and quality of BoNTs for pharmaceutical use.
Asunto(s)
Toxinas Botulínicas Tipo A/biosíntesis , Toxinas Botulínicas Tipo A/genética , Clostridium botulinum/genética , Clostridium botulinum/metabolismo , Regulación de la Expresión Génica , Procesamiento Proteico-Postraduccional/genética , Animales , Arginina/metabolismo , Arginina/farmacología , Clostridium botulinum/efectos de los fármacos , Medios de Cultivo/química , Femenino , Concentración de Iones de Hidrógeno , Ratones , Ratones Endogámicos ICRRESUMEN
Outgrowth and toxinogenesis of Clostridium botulinum Group II (non-proteolytic) type B were studied in cooked ham prepared with different NaNO2 (ranging from 0 to 80â¯mg/kg) and sodium chloride (NaCl, ranging from 12 to 19â¯g/kg) incorporation rates. Cured ground pork batters were inoculated with a cocktail of 3 strains of C. botulinum Group II type B at 3.5 log10 CFU/g, portioned and samples of 50â¯g were vacuum packed then cooked and cooled based on thermal processing employed by the meat processing industry. These cooked ham model samples were stored under reasonably foreseeable conditions of use and storage i.e. for 14â¯days at 4⯰C, followed by a cold chain break for 1â¯h at 20⯰C then up to 33â¯days at 8⯰C. Storage times and temperatures were used to mimic those commonly encountered along the supply chain. Enumeration of C. botulinum and detection of the botulinum neurotoxin type B (BoNT/B) were performed in triplicate at different storage times. Under these experimental conditions, incorporation rates of NaNO2â¯≥â¯30â¯mg/kg prevented the outgrowth and toxinogenesis of C. botulinum Group II type B in the cooked ham model, regardless of the NaCl concentrations tested. In contrast, total removal of nitrite allowed outgrowth and toxin production during storage of the processed meat product. Results showed that the maximum ingoing amount of nitrite (i.e. 150â¯mg/kg) that may be added according to the EU legislation (Regulation (EC) No 1333/2008) can be reduced in cooked ham while still ensuring control of C. botulinum Group II type B. According to the multiple factors that could affect C. botulinum behavior in processing meat products, outgrowth and toxin production of C. botulinum should be evaluated on a case by case basis, depending on the recipe, manufacturing process, food matrix and storage conditions.
Asunto(s)
Clostridium botulinum/crecimiento & desarrollo , Clostridium botulinum/metabolismo , Conservantes de Alimentos/análisis , Carne de Cerdo/microbiología , Nitrito de Sodio/análisis , Animales , Toxinas Botulínicas/análisis , Toxinas Botulínicas/metabolismo , Clostridium botulinum/efectos de los fármacos , Frío , Recuento de Colonia Microbiana , Culinaria , Manipulación de Alimentos/métodos , Manipulación de Alimentos/normas , Conservantes de Alimentos/farmacología , Cloruro de Sodio/análisis , Cloruro de Sodio/farmacología , Nitrito de Sodio/farmacología , VacioRESUMEN
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.
Asunto(s)
Botulismo/diagnóstico , Antibacterianos/administración & dosificación , Botulismo/tratamiento farmacológico , Botulismo/epidemiología , Clostridium botulinum/efectos de los fármacos , Clostridium botulinum/patogenicidad , Trastornos de la Conciencia , Femenino , Fiebre/etiología , Hawaii/epidemiología , Humanos , Lactante , MasculinoRESUMEN
Botulinum neurotoxin (BoNT) is the most potent natural toxin known. Of the seven BoNT serotypes (A to G), types A, B, E, and F cause human botulism. Treatment of human botulism requires the development of effective toxin-neutralizing antibodies without side effects such as serum sickness and anaphylaxis. In this study, we generated fully human monoclonal antibodies (HuMAbs) against serotype B BoNT (BoNT/B1) using a murine-human chimera fusion partner cell line named SPYMEG. Of these HuMAbs, M2, which specifically binds to the light chain of BoNT/B1, showed neutralization activity in a mouse bioassay (approximately 10 i.p. LD50/100 µg of antibody), and M4, which binds to the C-terminal of heavy chain, showed partial protection. The combination of two HuMAbs, M2 (1.25 µg) and M4 (1.25 µg), was able to completely neutralize BoNT/B1 (80 i.p. LD50) with a potency greater than 80 i.p. LD50/2.5 µg of antibodies, and was effective both prophylactically and therapeutically in the mouse model of botulism. Moreover, this combination showed broad neutralization activity against three type B subtypes, namely BoNT/B1, BoNT/B2, and BoNT/B6. These data demonstrate that the combination of M2 and M4 is promising in terms of a foundation for new human therapeutics for BoNT/B intoxication.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Toxinas Botulínicas Tipo A/antagonistas & inhibidores , Botulismo/prevención & control , Anticuerpos ampliamente neutralizantes/farmacología , Clostridium botulinum/efectos de los fármacos , Animales , Anticuerpos Monoclonales/inmunología , Especificidad de Anticuerpos , Sitios de Unión de Anticuerpos , Toxinas Botulínicas Tipo A/inmunología , Botulismo/inmunología , Botulismo/microbiología , Anticuerpos ampliamente neutralizantes/inmunología , Clostridium botulinum/inmunología , Modelos Animales de Enfermedad , Quimioterapia Combinada , Epítopos , Femenino , Humanos , Hibridomas , Ratones , Pruebas de Neutralización , Unión ProteicaRESUMEN
Botulinum neurotoxins (BoNTs) are the most toxic category A biological warfare agents. There is no therapeutics available for BoNT intoxication yet, necessitating the development of a medical countermeasure against these neurotoxins. The discovery of small molecule-based drugs has revolutionized in the last two decades resulting in the identification of several small molecule inhibitors of BoNTs. However, none progressed to clinical trials. 8-Hydroxyquinolines scaffold-based molecules are important 'privileged structures' that can be exploited as inhibitors of a diverse range of targets. In this review, our study of recent reports suggests the development of 8-hydroxyquinoline derived molecules as a potential drug may be on the horizon.
Asunto(s)
Neurotoxinas/antagonistas & inhibidores , Oxiquinolina/farmacología , Bibliotecas de Moléculas Pequeñas/farmacología , Animales , Clostridium botulinum/química , Clostridium botulinum/efectos de los fármacos , Humanos , Estructura Molecular , Oxiquinolina/química , Bibliotecas de Moléculas Pequeñas/químicaRESUMEN
Growing antimicrobial resistance of the pathogens against multiple drugs posed a serious threat to the human health worldwide. This fueled the need of identifying the novel therapeutic targets that can be used for developing new class of the drugs. Recently, there is a substantial rise in the rate of Clostridium infections as well as in the emergence of virulent and antibiotic resistant strains. Hence, there is an urgent need for the identification of potential therapeutic targets and the development of new drugs for the treatment and prevention of Clostridium infections. In the present study, a combinatorial approach involving systems biology and comparative genomics strategy was tested against Clostridium botulinum ATCC 3502 and Clostridium difficile str. 630 pathogens, to render potential therapeutic target at qualitative and quantitative level. This resulted in the identification of five common (present in both the pathogens, 34 in C. botulinum ATCC 3502 and 42 in C. difficile str. 630) drug targets followed by virtual screening-based identification of potential inhibitors employing molecular docking and molecular dynamics simulations. The identified targets will provide a solid platform for the designing of novel wide-spectrum lead compounds capable of inhibiting their catalytic activities against multidrug-resistant Clostridium in the near future.
Asunto(s)
Antibacterianos/farmacología , Clostridioides difficile/patogenicidad , Clostridium botulinum/patogenicidad , Proteómica/métodos , Factores de Virulencia/metabolismo , Proteínas Bacterianas/efectos de los fármacos , Proteínas Bacterianas/metabolismo , Clostridioides difficile/efectos de los fármacos , Clostridioides difficile/metabolismo , Clostridium botulinum/efectos de los fármacos , Clostridium botulinum/metabolismo , Simulación por Computador , Diseño de Fármacos , Farmacorresistencia Bacteriana/efectos de los fármacos , Genómica , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Biología de SistemasRESUMEN
Apesar da utilização dos aditivos nos alimentos, alguns microrganismos, como o Clostridium botulinum ainda causam preocupação. Óleos essenciais são opções de aditivo para a indústria de alimentos, pois podem apresentar atividade antimicrobiana e antioxidante. Diante disso, o objetivo foi extrair o ó leo essencial de Lippia gracilis, caracterizar sua composição química por CG/ EM, avaliar a atividade antioxidante pela redução do radical DPPH e a concentração mínima esporicida (CME) sobreClostridium sporogenes. O óleo essencial é composto majoritariamente pelos monoterpenos carvacrol, p-cimeno e terpineno, e apresentou baixa capacidade de estabilização do radical DPPH, IC50 > 500 μg/ mL. Em relação à ação sobre os endósporos do Clostridium sporogenes, a concentração mínima esporicida foi 1,5% de óleo essencial.
Asunto(s)
Antiinfecciosos , Antioxidantes/análisis , Clostridium botulinum/efectos de los fármacos , Clostridium botulinum/patogenicidad , Lippia , Aceites Volátiles/análisis , Aceites Volátiles/farmacologíaRESUMEN
A forma mais comum de inibir a produção de toxina botulínica em produtos cárneos cozidos é pela adição de sais de nitrito, o que pode gerar substâncias carcinogênicas (nitrosaminas), sendo desejável sua substituição. Os óleos essenciais vêm se destacando como agentes antimicrobianos, sendo interessante seu uso como conservante. O objetivo desse trabalho foi avaliar a ação sinergística de óleos essenciais sobre endósporos de Clostridium sporogenes, utilizado como modelo de pesquisa para C. botulinum. As concentrações mínimas esporicidas (CME) dos óleos de alecrim, tomilho, cravo, manjericão, ho wood e alho foram de 3% e de 0,375% para pimenta e canela. Os óleos de orégano e noz moscada não apresentaram ação esporicida nas concentrações testadas. Entre as combinações, as melhores foram de pimenta chinesa (0,1306%), alho (1,1%) e manjericão (1,1%) e pimenta chinesa (0,1306%), alho (1,1%) e tomilho branco (1,1%). Os resultados sugerem o sinergismo entre os óleos, sendo promissor seu uso em alimentos.
Asunto(s)
Antiinfecciosos , Clostridium/efectos de los fármacos , Conservantes de Alimentos , Aceites Volátiles/administración & dosificación , Clostridium botulinum/efectos de los fármacos , Sinergismo FarmacológicoRESUMEN
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]).
Asunto(s)
Antibacterianos/farmacología , Botulismo/tratamiento farmacológico , Botulismo/microbiología , Clostridium botulinum/efectos de los fármacos , California , Clostridium/aislamiento & purificación , Clostridium/patogenicidad , Clostridium botulinum/aislamiento & purificación , Farmacorresistencia Bacteriana/efectos de los fármacos , Femenino , Humanos , Lactante , Pruebas de Sensibilidad MicrobianaRESUMEN
The bio molecules from plant leaf extract utilized in the preparation of selenium material at the nano scale. The selenium ion was reduced to selenium nanoparticles in the presence of molecule residue of the plant leaf extract. The bio molecule stabilized selenium nanoparticles were grown gradually in the reaction mixture. The selenium nanoparticles were characterized using atomic absorption spectroscopy, fourier transform inferred spectroscopy, X-ray diffraction, scanning electronic microscope and transmission electronic microscope. The selenium nanoparticles were synthesized successfully as the nano-crystalline pure hexagonal phase and the size range of 26-41â¯nm with spherical in shape. The activity and mechanism of nanoparticles suggested that the selenium nanoparticles are causes of leakage of reducing sugars and protein of pathogens membrane cell. The selenium nano are responsible for death and fully inhibited the microbial growth of pathogen. The bio molecule stabilized selenium nanoparticles were also investigated for the antioxidant agent. Selenium nanoparticles showed scavenging activity up to 94.48%. These results recommended that the advantages of using this method for synthesis of selenium nanoparticles with excellent antioxidant and antimicrobial mechanism and activity, which can be used as the antioxidant and antibiotic agent.
Asunto(s)
Antibacterianos/farmacología , Clostridium botulinum/efectos de los fármacos , Nanopartículas del Metal/química , Oxidación-Reducción/efectos de los fármacos , Extractos Vegetales/farmacología , Selenio/farmacología , Antioxidantes/farmacología , Azadirachta/química , Membrana Celular/efectos de los fármacos , Clostridium botulinum/crecimiento & desarrollo , Nanopartículas del Metal/ultraestructura , Pruebas de Sensibilidad Microbiana , Tamaño de la Partícula , Hojas de la Planta/químicaRESUMEN
The 1986 Food Research Institute-Tanaka et al. model predicts the safety of shelf-stable process cheese spread formulations using the parameters of moisture, pH, NaCl, and disodium phosphate (DSP) to inhibit toxin production by Clostridium botulinum. Although this model is very reliable for predicting safety for standard-of-identity spreads, the effects of additional factors have not been considered. The objective of this study was to create a predictive model to include the interactive effect of moisture, pH, fat, sorbic acid, and potassium-based replacements for NaCl and DSP to reflect modern reduced-sodium recipes. Eighty formulations were identified using a central composite design targeting seven factors: 50 to 60% moisture, pH 5.4 to 6.2, 0 to 0.2% sorbic acid, 10 to 30% fat, 1.7 to 2.4% NaCl, 0.8 to 1.6% DSP, and 0 to 50% potassium replacement for sodium salts. Samples were inoculated with proteolytic C. botulinum spores at 3 log spores per g, hot filled into sterile vials, and stored anaerobically at 27°C. Samples were assayed at 0, 1, 2, 3, 4, 8.5, 17.5, 26, and 40 weeks for the presence of botulinum toxin using the mouse bioassay. A parametric survival model was fit to the censored time-to-toxin data. All linear, quadratic, and pairwise effects were considered for model fit. As hypothesized, the effects of pH, sorbate, moisture, DSP, and NaCl were highly significant (P < 0.001). Fat concentration and potassium replacement effects were significant at P < 0.021 and P < 0.057, respectively. The model consistently predicted the safety failure of the toxic samples, but it also predicted failure for some samples that were not toxic. This model is an adjunct to existing models by adding the factors of potassium salts, fat, and sorbic acid to predict the botulinal safety of prepared process cheese products but is not intended to be a substitute for formulation evaluation by a competent process authority.
Asunto(s)
Toxinas Botulínicas/biosíntesis , Queso/microbiología , Clostridium botulinum/crecimiento & desarrollo , Microbiología de Alimentos , Conservación de Alimentos/métodos , Animales , Clostridium botulinum/efectos de los fármacos , Clostridium botulinum/metabolismo , Seguridad de Productos para el Consumidor , Humanos , Concentración de Iones de Hidrógeno , Sodio , TemperaturaRESUMEN
Clostridium botulinum, a formidable pathogen is responsible for the emerging cause of food poisoning cases on the global canvas. The endemicity of bacterium Clostridium botulinum is reflected by the sudden hospital outbreaks and increased resistance towards multiple drugs. Therefore, a combined approach of in-silico comparative genomic analysis with statistical analysis was applied to overcome the limitation of bench-top technologies. Owing to the paucity of genomic data available by the advent of third generation sequencing technologies, several 'omics' technologies were applied to understand the underlying evolutionary pattern and lifestyle of the bacterial pathogen using phylogenomics. The calculation of pan-genome, core genome and singletons provides view of genetic repertoire of the bacterial pathogen lineage at the successive level, orthology shared and specific gene subsets. In addition, assessment of pathogenomic potential, resistome, toxin/antitoxin family in successive pathogenic strains of Clostridium botulinum aids in revealing more specific targets for drug design and development.
Asunto(s)
Clostridium botulinum/genética , Farmacorresistencia Bacteriana/genética , Genoma Bacteriano , Clostridium botulinum/clasificación , Clostridium botulinum/efectos de los fármacos , Clostridium botulinum/patogenicidad , Filogenia , Virulencia/genéticaAsunto(s)
Botulismo/tratamiento farmacológico , Inmunoglobulinas/administración & dosificación , Antitoxina Botulínica/administración & dosificación , Botulismo/diagnóstico , Botulismo/fisiopatología , Clostridium botulinum/efectos de los fármacos , Clostridium botulinum/aislamiento & purificación , Femenino , Humanos , Lactante , Resultado del TratamientoRESUMEN
The present study aimed to characterize Enterococcus faecalis (n = -6) and Enterococcus faecium (n = 1) isolated from healthy chickens to find a novel perspective probiotic candidate that antagonize Clostridium botulinum types A, B, D, and E. The isolated enterococci were characterized based on phenotypic properties, PCR, and matrix-assisted laser desorption/ionization time of flight (MALDI-TOF). The virulence determinants including hemolytic activity on blood agar, gelatinase activity, sensitivity to vancomycin, and presence of gelatinase (gelE) and enterococcal surface protein (esp) virulence genes were investigated. Also, the presence of enterocin structural genes enterocin A, enterocin B, enterocin P, enterocin L50A/B, bacteriocin 31, enterocin AS48, enterocin 1071A/1071B, and enterocin 96 were assessed using PCR. Lastly, the antagonistic effect of the selected Enterococcus spp. on the growth of C. botulinum types A, B, D, and E was studied. The obtained results showed that four out of six E. faecalis and one E. faecium proved to be free from the tested virulence markers. All tested enterococci strains exhibited more than one of the tested enterocin. Interestingly, E. faecalis and E. faecium significantly restrained the growth of C. botulinum types A, B, D, and E. In conclusion, although, the data presented showed that bacteriocinogenic Enterococcus strains lacking of virulence determinants could be potentially used as a probiotic candidate against C. botulinum in vitro; however, further investigations are still urgently required to verify the beneficial effects of the tested Enterococcus spp. in vivo.
Asunto(s)
Bacteriocinas/metabolismo , Clostridium botulinum/efectos de los fármacos , Enterococcus faecalis/genética , Enterococcus faecium/genética , Probióticos/farmacología , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Bacteriocinas/genética , Pollos/microbiología , Clostridium botulinum/crecimiento & desarrollo , Enterococcus faecalis/aislamiento & purificación , Enterococcus faecalis/metabolismo , Enterococcus faecium/aislamiento & purificación , Enterococcus faecium/metabolismo , Genotipo , Fenotipo , Probióticos/metabolismoRESUMEN
Botulinum neurotoxins (BoNTs) cause botulism and are the deadliest naturally-occurring substances known to humans. BoNTs have been classified as one of the category A agents by the Centers for Disease Control and Prevention, indicating their potential use as bioweapons. To counter bio-threat and naturally-occurring botulism cases, well-tolerated antibodies by humans that neutralize BoNTs are relevant. In our previous work, we showed the neutralizing potential of macaque (Macaca fascicularis)-derived scFv-Fc (scFv-Fc ELC18) by in vitro endopeptidase immunoassay and ex vivo mouse phrenic nerve-hemidiaphragm assay by targeting the light chain of the botulinum neurotoxin type E (BoNT/E). In the present study, we germline-humanized scFv-Fc ELC18 into a full IgG hu8ELC18 to increase its immunotolerance by humans. We demonstrated the protection and prophylaxis capacity of hu8ELC18 against BoNT/E in a mouse model. A concentration of 2.5 ng/mouse of hu8ELC18 protected against 5 mouse lethal dose (MLD) in a mouse protection assay and complete neutralization of 1 LD50 of pure BoNT/E toxin was achieved with 8 ng of hu8ELC18 in mouse paralysis assay. Furthermore, hu8ELC18 protected mice from 5 MLD if injected up to 14 days prior to intraperitoneal BoNT/E administration. This newly-developed humanized IgG is expected to have high tolerance in humans.
Asunto(s)
Anticuerpos Monoclonales Humanizados/farmacología , Anticuerpos Neutralizantes/farmacología , Antídotos/farmacología , Antitoxinas/farmacología , Toxinas Botulínicas/antagonistas & inhibidores , Botulismo/prevención & control , Clostridium botulinum/efectos de los fármacos , Anticuerpos de Cadena Única/farmacología , Animales , Toxinas Botulínicas/inmunología , Botulismo/inmunología , Botulismo/microbiología , Clostridium botulinum/inmunología , Clostridium botulinum/metabolismo , Modelos Animales de Enfermedad , Femenino , RatonesRESUMEN
Botulinum neurotoxins (BoNTs) are the most poisonous biological substance known to humans. They cause flaccid paralysis by blocking the release of acetylcholine at the neuromuscular junction. Here, we report a number of small molecule non-peptide inhibitors of BoNT serotype E. The structure-activity relationship and a pharmacophore model are presented. Although non-peptidic in nature, these inhibitors mimic key features of the uncleavable substrate peptide Arg-Ile-Met-Glu (RIME) of the SNAP-25 protein. Among the compounds tested, most of the potent inhibitors bear a zinc-chelating moiety connected to a hydrophobic and aromatic moiety through a carboxyl or amide linker. All of them show low micromolar IC50 values.