scholarly journals Integration of Complete Plasmids Containing Bont Genes into Chromosomes of Clostridium parabotulinum, Clostridium sporogenes, and Clostridium argentinense

Toxins ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 473
Author(s):  
Theresa J. Smith ◽  
Renmao Tian ◽  
Behzad Imanian ◽  
Charles H. D. Williamson ◽  
Shannon L. Johnson ◽  
...  
Keyword(s):  
Chromosome Analysis ◽  
Gene Clusters ◽  
Clostridium Sporogenes ◽  
Bacterial Proteins ◽  
Extrachromosomal Elements ◽  
Heterogenous Group ◽  
Key Factor ◽  
Botulinum Neurotoxins ◽  
Clostridial Species ◽  
Genomic Locations

At least 40 toxin subtypes of botulinum neurotoxins (BoNTs), a heterogenous group of bacterial proteins, are produced by seven different clostridial species. A key factor that drives the diversity of neurotoxigenic clostridia is the association of bont gene clusters with various genomic locations including plasmids, phages and the chromosome. Analysis of Clostridium sporogenes BoNT/B1 strain CDC 1632, C. argentinense BoNT/G strain CDC 2741, and Clostridium parabotulinum BoNT/B1 strain DFPST0006 genomes revealed bont gene clusters within plasmid-like sequences within the chromosome or nested in large contigs, with no evidence of extrachromosomal elements. A nucleotide sequence (255,474 bp) identified in CDC 1632 shared 99.5% identity (88% coverage) with bont/B1-containing plasmid pNPD7 of C. sporogenes CDC 67071; CDC 2741 contig AYSO01000020 (1.1 MB) contained a ~140 kb region which shared 99.99% identity (100% coverage) with plasmid pRSJ17_1 of C. argentinense BoNT/G strain 89G; and DFPST0006 contig JACBDK0100002 (573 kb) contained a region that shared 100% identity (99%) coverage with the bont/B1-containing plasmid pCLD of C. parabotulinum Okra. This is the first report of full-length plasmid DNA-carrying complete neurotoxin gene clusters integrated in three distinct neurotoxigenic species: C. parabotulinum, C. sporogenes and C. argentinense.

scholarly journals Fimbrial ghf Gene Cluster of Genital Strains of Haemophilus spp.

2002 ◽  
Vol 70 (10) ◽  
pp. 5438-5445 ◽  
Author(s):  
Guillaume Bruant ◽  
Nathalie Gousset ◽  
Roland Quentin ◽  
Agnes Rosenau
Keyword(s):  
Gene Cluster ◽  
Chromosome Analysis ◽  
Gene Clusters ◽  
Large Deletion ◽  
Gene Promoters ◽  
Rt Pcr ◽  
Divergent Promoters ◽  
Intergenic Regions ◽  
Flanking Regions ◽  
Palindromic Sequences

ABSTRACT We analyzed the LKP fimbrial gene clusters of six piliated strains of a cryptic genospecies of Haemophilus isolated from the genital tracts of adult patients (five strains) and from an infected neonate. In a group of 19 genital strains, LKP-like genes have been found in only these 6 strains. In addition to the ghfA, ghfD, and ghfE genes previously described, we characterized two genes, designated ghfB and ghfC, encoding the putative chaperone and assembly platform proteins. All six strains had a complete and unique LKP-like gene cluster consisting of the five genes ghfA to ghfE, homologous to genes hifA to hifE of Haemophilus influenzae. The sequences of the coding and intergenic regions of the ghf clusters of the six strains were remarkably homologous. Unlike hif clusters, which are inserted between purE and pepN, the ghf cluster was inserted between purK and pepN on the chromosome. Analysis of the flanking regions of the ghf cluster identified a large deletion, identical in the 5′ end regions of all strains, including the whole purE gene and much of the purK gene. Ultrastructural observations, an attempt at enriching LKP fimbriae, and hemagglutination experiments demonstrated that none of the strains had LKP-type fimbriae. Nevertheless, reverse transcription (RT)-PCR showed that ghf genes were transcribed in four of the six strains. Sequencing of the intergenic ghfA-ghfB regions, including the ghf gene promoters, showed that the absence of transcripts in the remaining two strains was due to a decrease in the number of TA repeats (4 or 9 repeats rather than 10) between the −10 and −35 boxes of the two overlapping and divergent promoters. The other four strains, which had ghf transcripts, had the optimal 10 TA repeats (one strain) or 5 repeats associated with putative alternative −35 boxes (three strains). The absence of 10 repeated palindromic sequences of 44 or 45 nucleotides upstream of ghfB induces an increased instability of mRNA, as quantified by real-time RT-PCR, and may explain why the LKP fimbrial gene cluster is not expressed in these strains.

scholarly journals Methanosarcina acetivorans simultaneously produces molybdenum, vanadium, and iron-only nitrogenases in response to fixed nitrogen and molybdenum depletion

2021 ◽  
Author(s):  
Melissa Chanderban ◽  
Christopher A Hill ◽  
Ahmed E Dhamad ◽  
Daniel J Lessner
Keyword(s):  
Growth Rate ◽  
Gene Clusters ◽  
Immunoblot Analysis ◽  
Cell Yield ◽  
Methanosarcina Acetivorans ◽  
Fixed Nitrogen ◽  
Qpcr Analysis ◽  
Key Factor ◽  
Alternative Nitrogenase ◽  
Diazotrophic Growth

All nitrogen-fixing bacteria and archaea (diazotrophs) use molybdenum (Mo) nitrogenase to reduce dinitrogen (N2) to ammonia. Some diazotrophs also contain alternative nitrogenases that lack Mo: vanadium (V) and iron-only (Fe) nitrogenases. Among diazotrophs, the regulation and usage of the alternative nitrogenases in methanogens is largely unknown. Methanosarcina acetivorans contains nif, vnf, and anf gene clusters encoding putative Mo-, V-, and Fe-nitrogenases, respectively. This study investigated the effect of fixed nitrogen and Mo/V availability on nitrogenase expression and growth by M. acetivorans. The availability of Mo and V did not affect growth of M. acetivorans with fixed nitrogen but significantly affected growth with N2. M. acetivorans exhibited the fastest growth rate and highest cell yield during growth with N2 in medium containing Mo. Depletion of Mo (Fe-only condition) resulted in a significant decrease in growth rate and cell yield. The addition of V to Mo-depleted medium stimulated diazotrophic growth but was still less than growth in Mo-replete medium. qPCR analysis revealed transcription of the nif operon is only moderately affected by depletion of fixed nitrogen and Mo. However, vnf and anf transcription increased significantly when fixed nitrogen and Mo were depleted, with removal of Mo being the key factor. Immunoblot analysis revealed Mo-nitrogenase is produced when fixed nitrogen is depleted regardless of Mo availability, while V- and Fe-nitrogenases are produced only in the absence of fixed nitrogen and Mo. These results reveal that alternative nitrogenase production in M. acetivorans is tightly controlled and that all three nitrogenases can be simultaneously produced.

scholarly journals Unexpected Micro-Spatial Scale Genomic Diversity of the Bloom-Forming Cyanobacterium Aphanizomenon Gracile and its Phycosphere

2021 ◽  
Author(s):  
Sebastien Halary ◽  
Sébastien Duperron ◽  
Sandra Kim Tiam ◽  
Charlotte Duval ◽  
Eloïse Dhénain ◽  
...  
Keyword(s):  
Heterotrophic Bacteria ◽  
Hot Spot ◽  
Genotypic Diversity ◽  
Gene Clusters ◽  
Genomic Diversity ◽  
Key Factor ◽  
Colonization Success ◽  
Spatio Temporal ◽  
Water Ecosystems ◽  
Aphanizomenon Gracile

Abstract Background: Genotypic diversity within cyanobacteria populations, partly driven by horizontal gene transfers, is a key factor of their colonization success, allowing them to cope with spatio-temporal fluctuations of the environmental conditions. By providing complementary functions, such as oxidative stress protection, heterotrophic bacteria composing phycospheres play also an essential role in cyanobacteria adaptation. Aphanizomenon gracile is a species of toxinogen cyanobacteria blooming worldwide with severe consequences for fresh and brackish water ecosystems. While marker heterogeneity surveys have shown that harmful cyanobacteria blooms were not clonal populations, the real extent of genetic and functional diversity within a population of freshwater cyanobacteria, including A. gracile , and their associated phycospheres remains unclear. Results: Here, comparative omics of four monoclonal strains of A. gracile isolated from a single drop of water reveals extensive heterogeneity of chemotypes and gene contents, despite constrained genome size and high similarity indices. These variations are remarkably associated with horizontal gene transfers (HGT) of biosynthetic gene clusters (BCG), and a novel siphophage infecting A. gracile displaying characteristics of temperate phages appears to participate to this genotypic diversification. In spite of high variability in heterotrophic taxa relative abundances, A. gracile phycospheres displayed an apparent functional redundancy implying biosynthesis of public goods. Conclusions: Altogether, these results suggest that a bloom would constitute a hot-spot for A. gracile genotype diversification driven by cyanophages, where losses and gains of BCGs compels cyanobacteria individuals to cooperate together and with heterotrophic bacteria in a black queen hypothesis compatible way.

scholarly journals Botulinum Neurotoxin-Producing Bacteria. Isn’t It Time that We Called a Species a Species?

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Theresa Smith ◽  
Charles H. D. Williamson ◽  
Karen Hill ◽  
Jason Sahl ◽  
Paul Keim
Keyword(s):  
Clostridium Botulinum ◽  
Genomic Analysis ◽  
Single Species ◽  
Species Group ◽  
Secondary System ◽  
Content Type ◽  
Botulinum Neurotoxins ◽  
Clostridial Species ◽  
Taxonomic Groups ◽  
Species Taxonomy

ABSTRACTBotulinum neurotoxins (BoNTs) are produced by a diverse set of seven clostridial species, though alternate naming systems have developed over the last 100 years. Starting in the 1950s, a single-species taxonomy where any bacterium producing BoNT would be designatedClostridium botulinumwas introduced. As the extreme diversity of these strains was recognized, a secondary system of taxonomic “groups” evolved. It became clear that these groups also had members that did not produce BoNT, and in some cases, they were given formal species names. Genomic analysis now clearly identifies species affiliations whether an isolate is toxigenic or not. It is clear thatC. botulinumgroup nomenclature is no longer appropriate and that there are recognized species names for each clostridium. We advocate for the use of the scientific binomials and that the single-species group nomenclature be abandoned.

scholarly journals Transcriptome and Co-Expression Network Analyses Identify the Molecular Signatures Underlying Drought Resistance in Yellowhorn

Forests ◽  
2020 ◽  
Vol 11 (8) ◽  
pp. 840 ◽  
Author(s):  
Xiaojuan Liu ◽  
Yifan Cui ◽  
Zhiyan Wu ◽  
Yang Zhao ◽  
Xiaoyu Hu ◽  
...  
Keyword(s):  
Drought Resistance ◽  
Molecular Mechanisms ◽  
Gene Clusters ◽  
Growth And Yield ◽  
Biological Processes ◽  
Hub Genes ◽  
Network Analyses ◽  
Drought Conditions ◽  
Key Factor ◽  
Use Efficiency

Drought is a key factor that limits plant growth and yield. Yellowhorn is an important and vigorously promoted oil tree in China. It can survive under certain drought conditions, but a lack of water severely restricts its growth and results in yield losses in arid and semi-arid areas. Therefore, it is important to identify the key pathways and genes to understand the mechanisms of its drought resistance. In this study, we evaluated drought resistance in four types of yellowhorn, and obtained 2669 and 2451 differentially expressed genes (DEGs) via the transcriptome analysis of the comparison of water-saving/water-consuming and fast-growing/slow-growing yellowhorn, respectively, under long-term drought conditions. The gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of DEGs showed the key biological processes and metabolic pathways involved in drought resistance, which demonstrated that there are both the same and different biological processes involved in regulating water use efficiency (WUE) and growth in response to drought stress. Furthermore, the network analysis indicated hub genes (especially seven protein kinases) and potential co-expressed gene clusters in a greenyellow module associated with WUE and a blue module associated with growth. These identified hub genes and key biological processes can significantly enhance our knowledge about the molecular mechanisms of drought resistance in yellowhorn.

scholarly journals Dynamin Inhibition Blocks Botulinum Neurotoxin Type A Endocytosis in Neurons and Delays Botulism

2011 ◽  
Vol 286 (41) ◽  
pp. 35966-35976 ◽  
Author(s):  
Callista B. Harper ◽  
Sally Martin ◽  
Tam H. Nguyen ◽  
Shari J. Daniels ◽  
Nickolas A. Lavidis ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Motor Nerve ◽  
Nerve Terminals ◽  
Dependent Manner ◽  
The Novel ◽  
Motor Nerve Terminals ◽  
Presynaptic Nerve Terminal ◽  
Bacterial Proteins ◽  
Fluorescence And Electron Microscopy ◽  
Botulinum Neurotoxins

The botulinum neurotoxins (BoNTs) are di-chain bacterial proteins responsible for the paralytic disease botulism. Following binding to the plasma membrane of cholinergic motor nerve terminals, BoNTs are internalized into an endocytic compartment. Although several endocytic pathways have been characterized in neurons, the molecular mechanism underpinning the uptake of BoNTs at the presynaptic nerve terminal is still unclear. Here, a recombinant BoNT/A heavy chain binding domain (Hc) was used to unravel the internalization pathway by fluorescence and electron microscopy. BoNT/A-Hc initially enters cultured hippocampal neurons in an activity-dependent manner into synaptic vesicles and clathrin-coated vesicles before also entering endosomal structures and multivesicular bodies. We found that inhibiting dynamin with the novel potent Dynasore analog, Dyngo-4aTM, was sufficient to abolish BoNT/A-Hc internalization and BoNT/A-induced SNAP25 cleavage in hippocampal neurons. Dyngo-4a also interfered with BoNT/A-Hc internalization into motor nerve terminals. Furthermore, Dyngo-4a afforded protection against BoNT/A-induced paralysis at the rat hemidiaphragm. A significant delay of >30% in the onset of botulism was observed in mice injected with Dyngo-4a. Dynamin inhibition therefore provides a therapeutic avenue for the treatment of botulism and other diseases caused by pathogens sharing dynamin-dependent uptake mechanisms.

Production of indole-3-propanoic acid and 3-(p-hydroxyphenyl)propanoic acid by Clostridium sporogenes: a convenient thin-layer chromatography detection system

1980 ◽  
Vol 26 (4) ◽  
pp. 448-453 ◽  
Author(s):  
Joanne J. Jellet ◽  
Thomas P. Forrest ◽  
Ian. A. Macdonald ◽  
Thomas J. Marrie ◽  
Lillian V. Holdeman
Keyword(s):  
Amino Acids ◽  
Thin Layer ◽  
Thin Layer Chromatography ◽  
Detection System ◽  
Spot Test ◽  
The Other ◽  
Propanoic Acid ◽  
Clostridium Sporogenes ◽  
Clostridial Species ◽  
Layer Chromatography

Indole-3-propanoic acid (IPA), 3-(p-hydroxyphenyl)propanoic acid (HPPA), and 3-phenylpropanoic acid (PPA) were present in the spent bacterial media of Clostridium sporogenes (20/20 strains) and C. cylindrosporum (1/1 strains), but absent in 32 other clostridial species (74 strains) tested. Both IPA and HPPA (but not PPA) could be readily detected by thin-layer chromatography and p-hydroxybenzaldehyde spray reagent. IPA forms a scarlet complex with p-hydroxybenzaldehyde which shifts to purple and remains stable for up to 6 weeks. IPA can be detected in acidified extracts of C. sporogenes by a simple spot test. The structures of IPA and HPPA were confirmed by nuclear magnetic resonance (nmr) and mass spectroscopy and their formation was detected by the absorbance at 280 nm. Addition of one of the precursor amino acids (L-tryptophan, L-tyrosine, or L-phenylalanine) to the medium greatly enhanced formation of the corresponding deaminated acid and depressed the formation of the other two acids. The products IPA, HPPA, and PPA, at 10−3M, and spent bacterial media were negative in the direct Ames's assay for mutagenicity and noncytotoxic towards MRC-S cells.

scholarly journals The Distinctive Evolution of orfX Clostridium parabotulinum Strains and Their Botulinum Neurotoxin Type A and F Gene Clusters Is Influenced by Environmental Factors and Gene Interactions via Mobile Genetic Elements

2021 ◽  
Vol 12 ◽  
Author(s):  
Theresa J. Smith ◽  
Charles H. D. Williamson ◽  
Karen K. Hill ◽  
Shannon L. Johnson ◽  
Gary Xie ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Botulinum Neurotoxin ◽  
Genetic Material ◽  
Gene Clusters ◽  
Toxin Gene ◽  
Diverse Range ◽  
Group I ◽  
Extrachromosomal Elements ◽  
Genomic Relationships ◽  
Recent Sequencing

Of the seven currently known botulinum neurotoxin-producing species of Clostridium, C. parabotulinum, or C. botulinum Group I, is the species associated with the majority of human botulism cases worldwide. Phylogenetic analysis of these bacteria reveals a diverse species with multiple genomic clades. The neurotoxins they produce are also diverse, with over 20 subtypes currently represented. The existence of different bont genes within very similar genomes and of the same bont genes/gene clusters within different bacterial variants/species indicates that they have evolved independently. The neurotoxin genes are associated with one of two toxin gene cluster types containing either hemagglutinin (ha) genes or orfX genes. These genes may be located within the chromosome or extrachromosomal elements such as large plasmids. Although BoNT-producing C parabotulinum bacteria are distributed globally, they are more ubiquitous in certain specific geographic regions. Notably, northern hemisphere strains primarily contain ha gene clusters while southern hemisphere strains have a preponderance of orfX gene clusters. OrfX C. parabotulinum strains constitute a subset of this species that contain highly conserved bont gene clusters having a diverse range of bont genes. While much has been written about strains with ha gene clusters, less attention has been devoted to those with orfX gene clusters. The recent sequencing of 28 orfX C. parabotulinum strains and the availability of an additional 91 strains for analysis provides an opportunity to compare genomic relationships and identify unique toxin gene cluster characteristics and locations within this species subset in depth. The mechanisms behind the independent processes of bacteria evolution and generation of toxin diversity are explored through the examination of bacterial relationships relating to source locations and evidence of horizontal transfer of genetic material among different bacterial variants, particularly concerning bont gene clusters. Analysis of the content and locations of the bont gene clusters offers insights into common mechanisms of genetic transfer, chromosomal integration, and development of diversity among these genes.

scholarly journals Botulinum-neurotoxin-like sequences identified from anEnterococcussp. genome assembly

2017 ◽  
Author(s):  
Charles H.D. Williamson ◽  
Theresa J. Smith ◽  
Brian T. Foley ◽  
Karen Hill ◽  
Paul Keim ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Genome Assembly ◽  
Botulinum Neurotoxin ◽  
Draft Genome ◽  
Gene Clusters ◽  
Flaccid Paralysis ◽  
Genomic Databases ◽  
Binding Motifs ◽  
Draft Genome Assembly ◽  
Botulinum Neurotoxins

AbstractBotulinum neurotoxins (BoNTs) are produced by diverse members of theClostridiaand result in a flaccid paralysis known as botulism. Exploring the diversity of BoNTs is important for the development of therapeutics and antitoxins. Here we describe a novel,bont-like gene cluster identified in a draft genome assembly forEnterococcussp. 3G1_DIV0629 by querying publicly available genomic databases. Thebont-like gene is found in a gene cluster similar to knownbontgene clusters. Protease and binding motifs conserved in known BoNT proteins are present in the newly identified BoNT-like protein; however, it is currently unknown if the BoNT-like protein described here is capable of targeting neuronal cells resulting in botulism.

scholarly journals Genomic Sequence and Characterization of the Virulent Bacteriophage φCTP1 from Clostridium tyrobutyricum and Heterologous Expression of Its Endolysin

2010 ◽  
Vol 76 (16) ◽  
pp. 5415-5422 ◽  
Author(s):  
Melinda J. Mayer ◽  
John Payne ◽  
Michael J. Gasson ◽  
Arjan Narbad
Keyword(s):  
Control Method ◽  
Genomic Sequence ◽  
Open Reading Frames ◽  
Clostridium Tyrobutyricum ◽  
Lytic Enzymes ◽  
Clostridium Sporogenes ◽  
E Coli ◽  
Virion Release ◽  
Gene Coding ◽  
Clostridial Species

ABSTRACT The growth of Clostridium tyrobutyricum in developing cheese leads to spoilage and cheese blowing. Bacteriophages or their specific lytic enzymes may provide a biological control method for eliminating such undesirable organisms without affecting other microflora. We isolated the virulent bacteriophage φCTP1 belonging to the Siphoviridae and have shown that it is effective in causing lysis of sensitive strains. The double-stranded DNA genome of φCTP1 is 59,199 bp, and sequence analysis indicated that it has 86 open reading frames. orf29 was identified as the gene coding for the phage endolysin responsible for cell wall degradation prior to virion release. We cloned and expressed the ctp1l gene in E. coli and demonstrated that the partially purified protein induced lysis of C. tyrobutyricum cells and reduced viable counts both in buffer and in milk. The endolysin was inactive against a range of clostridial species but did show lysis of Clostridium sporogenes, another potential spoilage organism. Removal of the C-terminal portion of the endolysin completely abolished lytic activity.

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