Differentiating acute bacterial infection from other causes of lower respiratory tract illness is challenging. In this trial, procalcitonin was investigated as a point-of-care test to aid in ...determining whether antibiotics were needed in the treatment of these illnesses.
The ability to form botulinum neurotoxin is restricted to six phylogenetically and physiologically distinct bacteria (Clostridium botulinum Groups I-IV and some strains of C. baratii and C. ...butyricum). The botulinum neurotoxin is the most potent toxin known, with as little as 30-100 ng potentially fatal, and is responsible for botulism, a severe neuroparalytic disease that affects humans, animals, and birds. In order to minimize the hazards presented by the botulinum neurotoxin-forming clostridia, it is necessary to extend understanding of the biology of these bacteria. Analyses of recently available genome sequences in conjunction with studies of bacterial physiology are beginning to reveal new and exciting information on the biology of these dangerous bacteria. At the whole organism level, substantial differences between the six botulinum neurotoxin-forming clostridia have been reported. For example, the genomes of proteolytic C. botulinum (C. botulinum Group I) and non-proteolytic C. botulinum (C. botulinum Group II) are highly diverged and show neither synteny nor homology. It has also emerged that the botulinum neurotoxin-forming clostridia are not overtly pathogenic (unlike C. difficile), but saprophytic bacteria that use the neurotoxin to kill a host and create a source of nutrients. One important feature that has contributed to the success of botulinum neurotoxin-forming clostridia is their ability to form highly resistant endospores. The spores, however, also present an opportunity to control these bacteria if escape from lag phase (and hence growth) can be prevented. This is dependent on extending understanding of the biology of these processes. Differences in the genetics and physiology of spore germination in proteolytic C. botulinum and non-proteolytic C. botulinum have been identified. The biological variability in lag phase and its stages has been described for individual spores, and it has been shown that various adverse treatments extend different stages of lag phase. For example, heat treatment primarily extended germination, while incubation at a chilled temperature primarily extended outgrowth. The neurotoxin gene is present within a cluster of associated genes, and can be located on the chromosome, a plasmid or a bacteriophage. Two basic types of neurotoxin cluster have been identified. Evolution of the neurotoxin gene and cluster has occurred independently of the organism, and involved a series of recombination events but is still poorly understood. Factors affecting the regulation of neurotoxin formation also remain poorly understood, and will be the focus of much future research.
Botulinum neurotoxins are diverse proteins. They are currently represented by at least seven serotypes and more than 40 subtypes. New clostridial strains that produce novel neurotoxin variants are ...being identified with increasing frequency, which presents challenges when organizing the nomenclature surrounding these neurotoxins. Worldwide, researchers are faced with the possibility that toxins having identical sequences may be given different designations or novel toxins having unique sequences may be given the same designations on publication. In order to minimize these problems, an ad hoc committee consisting of over 20 researchers in the field of botulinum neurotoxin research was convened to discuss the clarification of the issues involved in botulinum neurotoxin nomenclature. This publication presents a historical overview of the issues and provides guidelines for botulinum neurotoxin subtype nomenclature in the future.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Recent developments in whole genome sequencing have made a substantial contribution to understanding the genomes, neurotoxins and biology of Clostridium botulinum Group I (proteolytic C. botulinum) ...and C. botulinum Group II (non-proteolytic C. botulinum). Two different approaches are used to study genomics in these bacteria; comparative whole genome microarrays and direct comparison of complete genome DNA sequences. The properties of the different types of neurotoxin formed, and different neurotoxin gene clusters found in C. botulinum Groups I and II are explored. Specific examples of botulinum neurotoxin genes are chosen for an in-depth discussion of neurotoxin gene evolution. The most recent cases of foodborne botulism are summarised.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The deadly neurotoxins of Clostridium botulinum (BoNTs) comprise eight serotypes (A–G; X). The neurotoxin gene cluster encoding BoNT and its accessory proteins includes an operon containing an ntnh ...gene upstream of the boNT gene. Another operon contains either ha (haemagglutinin) or orfX genes (of unknown function). Here we describe a novel boNT gene cluster from Enterococcus sp. 3G1_DIV0629, with a typical ntnh gene and an uncommon orfX arrangement. The neurotoxin (designated putative eBoNT/J) contains a metallopeptidase zinc‐binding site, a translocation domain and a target cell attachment domain. Structural properties of the latter suggest a novel targeting mechanism with consequent implications for application by the pharmaceutical industry. This is the first complete boNT gene cluster identified in a non‐clostridial genome.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Lag phase represents the earliest and most poorly understood stage of the bacterial growth cycle. We developed a reproducible experimental system and conducted functional genomic and physiological ...analyses of a 2-h lag phase in Salmonella enterica serovar Typhimurium. Adaptation began within 4 min of inoculation into fresh LB medium with the transient expression of genes involved in phosphate uptake. The main lag-phase transcriptional program initiated at 20 min with the upregulation of 945 genes encoding processes such as transcription, translation, iron-sulfur protein assembly, nucleotide metabolism, LPS biosynthesis, and aerobic respiration. ChIP-chip revealed that RNA polymerase was not "poised" upstream of the bacterial genes that are rapidly induced at the beginning of lag phase, suggesting a mechanism that involves de novo partitioning of RNA polymerase to transcribe 522 bacterial genes within 4 min of leaving stationary phase. We used inductively coupled plasma mass spectrometry (ICP-MS) to discover that iron, calcium, and manganese are accumulated by S. Typhimurium during lag phase, while levels of cobalt, nickel, and sodium showed distinct growth-phase-specific patterns. The high concentration of iron during lag phase was associated with transient sensitivity to oxidative stress. The study of lag phase promises to identify the physiological and regulatory processes responsible for adaptation to new environments.
Spores of psychrotrophic Bacillus cereus may survive the mild heat treatments given to minimally processed chilled foods. Subsequent germination and cell multiplication during refrigerated storage ...may lead to bacterial concentrations that are hazardous to health.
This review is concerned with the characterisation of factors that prevent psychrotrophic B. cereus reaching hazardous concentrations in minimally processed chilled foods and associated foodborne illness. A risk assessment framework is used to quantify the risk associated with B. cereus and minimally processed chilled foods.
Bacillus cereus is responsible for two types of food poisoning, diarrhoeal (an infection) and emetic (an intoxication); however, no reported outbreaks of food poisoning have been associated with B. cereus and correctly stored commercially-produced minimally processed chilled foods. In the UK alone, more than 1010 packs of these foods have been sold in recent years without reported illness, thus the risk presented is very low. Further quantification of the risk is merited, and this requires additional data. The lack of association between diarrhoeal food poisoning and correctly stored commercially-produced minimally processed chilled foods indicates that an infectious dose has not been reached. This may reflect low pathogenicity of psychrotrophic strains. The lack of reported association of psychrotrophic B. cereus with emetic illness and correctly stored commercially-produced minimally processed chilled foods indicates that a toxic dose of the emetic toxin has not been formed. Laboratory studies show that strains form very small quantities of emetic toxin at chilled temperatures.
•Psychrotrophic B. cereus is a hazard to safety of minimally processed chilled foods.•Spores may survive food processing, with bacterial growth during chilled storage.•Illness not reported for B. cereus and correctly stored processed chilled foods.•Lack of foodborne illness probably due to low pathogenicity at chilled temperature.•High quality materials/manufacturing standards, and chill chain contribute to safety.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Clostridium botulinum is a dangerous pathogen that forms the highly potent botulinum toxin, which when ingested causes a deadly neuroparalytic disease. The closely related Clostridium sporogenes is ...occasionally pathogenic, frequently associated with food spoilage and regarded as the non-toxigenic equivalent of Group I C. botulinum. Both species form highly resistant spores that are ubiquitous in the environment and which, under favourable growth conditions germinate to produce vegetative cells. To improve the control of botulinum neurotoxin-forming clostridia, it is imperative to comprehend the mechanisms by which spores germinate. Germination is initiated following the recognition of small molecules (germinants) by a specific germinant receptor (GR) located in the spore inner membrane. The present study precisely defines clostridial GRs, germinants and co-germinants. Group I C. botulinum ATCC3502 contains two tricistronic and one pentacistronic GR operons, while C. sporogenes ATCC15579 has three tricistronic and one tetracistronic GR operons. Insertional knockout mutants, allied with characterisation of recombinant GRs shows for the first time that amino acid stimulated germination in C. botulinum requires two tri-cistronic encoded GRs which act in synergy and cannot function individually. Spore germination in C. sporogenes requires one tri-cistronic GR. Two other GRs form part of a complex involved in controlling the rate of amino-acid stimulated germination. The suitability of using C. sporogenes as a substitute for C. botulinum in germination studies and food challenge tests is discussed.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Proteolytic Clostridium botulinum is the causative agent of botulism, a severe neuroparalytic illness. Given the severity of botulism, surprisingly little is known of the population structure, ...biology, phylogeny or evolution of C. botulinum. The recent determination of the genome sequence of C. botulinum has allowed comparative genomic indexing using a DNA microarray.
Whole genome microarray analysis revealed that 63% of the coding sequences (CDSs) present in reference strain ATCC 3502 were common to all 61 widely-representative strains of proteolytic C. botulinum and the closely related C. sporogenes tested. This indicates a relatively stable genome. There was, however, evidence for recombination and genetic exchange, in particular within the neurotoxin gene and cluster (including transfer of neurotoxin genes to C. sporogenes), and the flagellar glycosylation island (FGI). These two loci appear to have evolved independently from each other, and from the remainder of the genetic complement. A number of strains were atypical; for example, while 10 out of 14 strains that formed type A1 toxin gave almost identical profiles in whole genome, neurotoxin cluster and FGI analyses, the other four strains showed divergent properties. Furthermore, a new neurotoxin sub-type (A5) has been discovered in strains from heroin-associated wound botulism cases. For the first time, differences in glycosylation profiles of the flagella could be linked to differences in the gene content of the FGI.
Proteolytic C. botulinum has a stable genome backbone containing specific regions of genetic heterogeneity. These include the neurotoxin gene cluster and the FGI, each having evolved independently of each other and the remainder of the genetic complement. Analysis of these genetic components provides a high degree of discrimination of strains of proteolytic C. botulinum, and is suitable for clinical and forensic investigations of botulism outbreaks.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Group I Clostridium botulinum and Clostridium sporogenes are physiologically and genetically closely related. Both are widely distributed in the environment and can cause foodborne botulism. In this ...work, a physiological study was conducted with 37 isolates from spoiled canned food and five referenced strains of C. sporogenes (three isolates) and Group I C. botulinum (two isolates). Growth limits of vegetative cells were established as a function of pH and NaCl concentration in PYG modified medium (PYGm) at 30 °C for 48 days. The heat resistance of the spores was studied for 2 min and 10 min at 102 °C and 110 °C. This physiological study (pH, NaCl growth limits and heat resistance) allowed the selection of 14 isolates of C. sporogenes (twelve isolates) and Group I C. botulinum (two isolates) representative of the diversity found. This panel of 14 selected isolates (11 isolated from spoiled canned food and three reference strains), were whole genome sequenced, but no association of physiological and genetic characteristics could be detected. Finally, we studied the ability of spores to germinate and grow from 5 isolates (four C. sporogenes and one Group I C. botulinum), under stress conditions generated by pH and NaCl following a low intensity heat treatment. The accumulation of these 3 stresses creates synergies that will strongly reduce the probability of spore growth in pH and salt conditions where they usually proliferate. The effect is progressive as the conditions become drastic: the number of decimal reduction observed increases translating a probability of growth which decreases. This study provides a better understanding of the behaviour of C. sporogenes and Group I C. botulinum isolates and shows how the combination of pH, NaCl and heat treatment can help prevent or minimise foodborne botulism outbreaks.
•Individual isolates of C. sporogenes and C. botulinum grew at pH ≥ 4.5 and NaCl ≤11.5%.•No correlation between physiological and genetic characteristics of 42 isolates.•pH and NaCl growth limits differed between vegetative cells and spores.•Combined low heat treatment, pH 4.9 and NaCl 4% prevented growth.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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