From Root to Tips: Sporulation Evolution and Specialization in Bacillus subtilis and the Intestinal Pathogen Clostridioides difficile

Abstract Bacteria of the Firmicutes phylum are able to enter a developmental pathway that culminates with the formation of highly resistant, dormant endospores. Endospores allow environmental persistence, dissemination and for pathogens, are also infection vehicles. In both the model Bacillus subtilis, an aerobic organism, and in the intestinal pathogen Clostridioides difficile, an obligate anaerobe, sporulation mobilizes hundreds of genes. Their expression is coordinated between the forespore and the mother cell, the two cells that participate in the process, and is kept in close register with the course of morphogenesis. The evolutionary mechanisms by which sporulation emerged and evolved in these two species, and more broadly across Firmicutes, remain largely unknown. Here, we trace the origin and evolution of sporulation using the genes known to be involved in the process in B. subtilis and C. difficile, and estimating their gain-loss dynamics in a comprehensive bacterial macroevolutionary framework. We show that sporulation evolution was driven by two major gene gain events, the first at the base of the Firmicutes and the second at the base of the B. subtilis group and within the Peptostreptococcaceae family, which includes C. difficile. We also show that early and late sporulation regulons have been coevolving and that sporulation genes entail greater innovation in B. subtilis with many Bacilli lineage-restricted genes. In contrast, C. difficile more often recruits new sporulation genes by horizontal gene transfer, which reflects both its highly mobile genome, the complexity of the gut microbiota, and an adjustment of sporulation to the gut ecosystem.

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From root to tips: sporulation evolution and specialization inBacillus subtilisand the intestinal pathogenClostridioides difficile

10.1101/473793 ◽

2019 ◽

Author(s):

Paula Ramos-Silva ◽

Mónica Serrano ◽

Adriano O. Henriques

Keyword(s):

Major Gene ◽

Gene Gain ◽

Environmental Persistence ◽

Evolutionary Mechanisms ◽

Origin And Evolution ◽

Clostridioides Difficile ◽

Intestinal Pathogen ◽

Dormant Spore ◽

Gain Loss ◽

Sporulation Genes

AbstractBacteria of the Firmicutes phylum are able to enter a developmental pathway that culminates with the formation of a highly resistant, dormant spore. Spores allow environmental persistence, dissemination and for pathogens, are infection vehicles. In both the modelBacillus subtilis, an aerobic species, and in the intestinal pathogenClostridioides difficile, an obligate anaerobe, sporulation mobilizes hundreds of genes. Their expression is coordinated between the forespore and the mother cell, the two cells that participate in the process, and is kept in close register with the course of morphogenesis. The evolutionary mechanisms by which sporulation emerged and evolved in these two species, and more broadly across Firmicutes, remain largely unknown. Here, we trace the origin and evolution of sporulation. Using the genes involved in the process inB. subtilisandC. difficile, and estimating their gain-loss dynamics in a comprehensive bacterial macro-evolutionary framework we show that sporulation evolution was driven by two major gene gain events, the first at the base of the Firmicutes and the second at the base of theB. subtilisgroup and within the Peptostreptococcaceae family, which includesC. difficile. We also show that early and late sporulation regulons have been co-evolving and that sporulation genes entail greater innovation inB. subtiliswith many Bacilli-lineage restricted genes. In contrast,C. difficilemore often recruits new sporulation genes by horizontal gene transfer, which reflects both its highly mobile genome, the complexity of the gut microbiota and an adjustment of sporulation to this particular ecosystem.

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Distribution and preservation of the components of the engulfment. What is beyond representative genomes?

PLoS ONE ◽

10.1371/journal.pone.0246651 ◽

2021 ◽

Vol 16 (3) ◽

pp. e0246651

Author(s):

Lizeth Soto-Avila ◽

Ricardo Ciria Merce ◽

Walter Santos ◽

Nori Castañeda ◽

Rosa-María Gutierrez-Ríos

Keyword(s):

Bacillus Subtilis ◽

Phylogenetic Reconstruction ◽

Leading Edge ◽

Extensive Study ◽

Origin And Evolution ◽

The Family ◽

Family Level ◽

Bioinformatic Approaches ◽

Gain Loss ◽

Synthesis And Degradation

Engulfment requires the coordinated, targeted synthesis and degradation of peptidoglycan at the leading edge of the engulfing membrane to allow the mother cell to completely engulf the forespore. Proteins such as the DMP and Q:AH complexes in Bacillus subtilis are essential for engulfment, as are a set of accessory proteins including GerM and SpoIIB, among others. Experimental and bioinformatic studies of these proteins in bacteria distinct from Bacillus subtilis indicate that fundamental differences exist regarding the organization and mechanisms used to successfully perform engulfment. As a consequence, the distribution and prevalence of the proteins involved in engulfment and other proteins that participate in different sporulation stages have been studied using bioinformatic approaches. These works are based on the prediction of orthologs in the genomes of representative Firmicutes and have been helpful in tracing hypotheses about the origin and evolution of sporulation genes, some of which have been postulated as sporulation signatures. To date, an extensive study of these signatures outside of the representative Firmicutes is not available. Here, we asked whether phyletic profiles of proteins involved in engulfment can be used as signatures able to describe the sporulation phenotype. We tested this hypothesis in a set of 954 Firmicutes, finding preserved phyletic profiles defining signatures at the genus level. Finally, a phylogenetic reconstruction based on non-redundant phyletic profiles at the family level shows the non-monophyletic origin of these proteins due to gain/loss events along the phylum Firmicutes.

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Role of SpoIVA ATPase Motifs during Clostridioides difficile Sporulation

Journal of Bacteriology ◽

10.1128/jb.00387-20 ◽

2020 ◽

Vol 202 (21) ◽

Author(s):

Hector Benito de la Puebla ◽

David Giacalone ◽

Alexei Cooper ◽

Aimee Shen

Keyword(s):

Bacillus Subtilis ◽

Atp Hydrolysis ◽

Spore Formation ◽

Spore Form ◽

Content Type ◽

Clostridioides Difficile ◽

Allele Specific ◽

Coat Layer ◽

Quality Control Mechanism

ABSTRACT The nosocomial pathogen Clostridioides difficile is a spore-forming obligate anaerobe that depends on its aerotolerant spore form to transmit infections. Functional spore formation depends on the assembly of a proteinaceous layer known as the coat around the developing spore. In C. difficile, coat assembly depends on the conserved spore protein SpoIVA and the clostridial-organism-specific spore protein SipL, which directly interact. Mutations that disrupt their interaction cause the coat to mislocalize and impair spore formation. In Bacillus subtilis, SpoIVA is an ATPase that uses ATP hydrolysis to drive its polymerization around the forespore. Loss of SpoIVA ATPase activity impairs B. subtilis SpoIVA encasement of the forespore and activates a quality control mechanism that eliminates these defective cells. Since this mechanism is lacking in C. difficile, we tested whether mutations in the C. difficile SpoIVA ATPase motifs impact functional spore formation. Disrupting C. difficile SpoIVA ATPase motifs resulted in phenotypes that were typically >104-fold less severe than the equivalent mutations in B. subtilis. Interestingly, mutation of ATPase motif residues predicted to abrogate SpoIVA binding to ATP decreased the SpoIVA-SipL interaction, whereas mutation of ATPase motif residues predicted to disrupt ATP hydrolysis but maintain ATP binding enhanced the SpoIVA-SipL interaction. When a sipL mutation known to reduce binding to SpoIVA was combined with a spoIVA mutation predicted to prevent SpoIVA binding to ATP, spore formation was severely exacerbated. Since this phenotype is allele specific, our data imply that SipL recognizes the ATP-bound form of SpoIVA and highlight the importance of this interaction for functional C. difficile spore formation. IMPORTANCE The major pathogen Clostridioides difficile depends on its spore form to transmit disease. However, the mechanism by which C. difficile assembles spores remains poorly characterized. We previously showed that binding between the spore morphogenetic proteins SpoIVA and SipL regulates assembly of the protective coat layer around the forespore. In this study, we determined that mutations in the C. difficile SpoIVA ATPase motifs result in relatively minor defects in spore formation, in contrast with Bacillus subtilis. Nevertheless, our data suggest that SipL preferentially recognizes the ATP-bound form of SpoIVA and identify a specific residue in the SipL C-terminal LysM domain that is critical for recognizing the ATP-bound form of SpoIVA. These findings advance our understanding of how SpoIVA-SipL interactions regulate C. difficile spore assembly.

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Life and Death of Selfish Genes: Comparative Genomics Reveals the Dynamic Evolution of Cytoplasmic Incompatibility

Molecular Biology and Evolution ◽

10.1093/molbev/msaa209 ◽

2020 ◽

Vol 38 (1) ◽

pp. 2-15 ◽

Cited By ~ 3

Author(s):

Julien Martinez ◽

Lisa Klasson ◽

John J Welch ◽

Francis M Jiggins

Keyword(s):

Cytoplasmic Incompatibility ◽

Gene Gain ◽

Evolutionary Models ◽

Loss Of Function ◽

Genome Sequences ◽

Life And Death ◽

Selfish Genes ◽

Wolbachia Genome ◽

The Cross ◽

Gain Loss

Abstract Cytoplasmic incompatibility is a selfish reproductive manipulation induced by the endosymbiont Wolbachia in arthropods. In males Wolbachia modifies sperm, leading to embryonic mortality in crosses with Wolbachia-free females. In females, Wolbachia rescues the cross and allows development to proceed normally. This provides a reproductive advantage to infected females, allowing the maternally transmitted symbiont to spread rapidly through host populations. We identified homologs of the genes underlying this phenotype, cifA and cifB, in 52 of 71 new and published Wolbachia genome sequences. They are strongly associated with cytoplasmic incompatibility. There are up to seven copies of the genes in each genome, and phylogenetic analysis shows that Wolbachia frequently acquires new copies due to pervasive horizontal transfer between strains. In many cases, the genes have subsequently acquired loss-of-function mutations to become pseudogenes. As predicted by theory, this tends to occur first in cifB, whose sole function is to modify sperm, and then in cifA, which is required to rescue the cross in females. Although cif genes recombine, recombination is largely restricted to closely related homologs. This is predicted under a model of coevolution between sperm modification and embryonic rescue, where recombination between distantly related pairs of genes would create a self-incompatible strain. Together, these patterns of gene gain, loss, and recombination support evolutionary models of cytoplasmic incompatibility.

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A Genomic Survey of Signalling in the Myxococcaceae

Microorganisms ◽

10.3390/microorganisms8111739 ◽

2020 ◽

Vol 8 (11) ◽

pp. 1739

Author(s):

David E. Whitworth ◽

Allison Zwarycz

Keyword(s):

Core Genome ◽

Gene Gain ◽

Fruiting Body Formation ◽

Two Component System ◽

Genome Sequences ◽

The Core ◽

Accessory Genes ◽

Two Component ◽

Gain Loss ◽

Core Genes

As prokaryotes diverge by evolution, essential ‘core’ genes required for conserved phenotypes are preferentially retained, while inessential ‘accessory’ genes are lost or diversify. We used the recently expanded number of myxobacterial genome sequences to investigate the conservation of their signalling proteins, focusing on two sister genera (Myxococcus and Corallococcus), and on a species within each genus (Myxococcus xanthus and Corallococcus exiguus). Four new C. exiguus genome sequences are also described here. Despite accessory genes accounting for substantial proportions of each myxobacterial genome, signalling proteins were found to be enriched in the core genome, with two-component system genes almost exclusively so. We also investigated the conservation of signalling proteins in three myxobacterial behaviours. The linear carotenogenesis pathway was entirely conserved, with no gene gain/loss observed. However, the modular fruiting body formation network was found to be evolutionarily plastic, with dispensable components in all modules (including components required for fruiting in the model myxobacterium M. xanthus DK1622). Quorum signalling (QS) is thought to be absent from most myxobacteria, however, they generally appear to be able to produce CAI-I (cholerae autoinducer-1), to sense other QS molecules, and to disrupt the QS of other organisms, potentially important abilities during predation of other prokaryotes.

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Systematic sequencing of the 283 kb 210 -232 region of the Bacillus subtilis genome containing the skin element and many sporulation genes

Microbiology ◽

10.1099/13500872-142-11-3103 ◽

1996 ◽

Vol 142 (11) ◽

pp. 3103-3111 ◽

Cited By ~ 30

Author(s):

M. Mizuno ◽

S. Masuda ◽

K.-i. Takemaru ◽

S. Hosono ◽

T. Sato ◽

...

Keyword(s):

Bacillus Subtilis ◽

Subtilis Genome ◽

Sporulation Genes

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Environmental Persistence of Bacillus anthracis and Bacillus subtilis Spores

PLoS ONE ◽

10.1371/journal.pone.0138083 ◽

2015 ◽

Vol 10 (9) ◽

pp. e0138083 ◽

Cited By ~ 18

Author(s):

Joseph P. Wood ◽

Kathryn M. Meyer ◽

Thomas J. Kelly ◽

Young W. Choi ◽

James V. Rogers ◽

...

Keyword(s):

Bacillus Subtilis ◽

Bacillus Anthracis ◽

Environmental Persistence

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Mutation in yaaT Leads to Significant Inhibition of Phosphorelay during Sporulation in Bacillus subtilis

Journal of Bacteriology ◽

10.1128/jb.184.20.5545-5553.2002 ◽

2002 ◽

Vol 184 (20) ◽

pp. 5545-5553 ◽

Cited By ~ 20

Author(s):

Shigeo Hosoya ◽

Kei Asai ◽

Naotake Ogasawara ◽

Michio Takeuchi ◽

Tsutomu Sato

Keyword(s):

Bacillus Subtilis ◽

Fluorescent Protein ◽

Early Stage ◽

Response Regulator ◽

Significant Inhibition ◽

Open Reading Frame ◽

Histidine Kinases ◽

Reading Frame ◽

Green Fluorescent ◽

Sporulation Genes

ABSTRACT In the course of a Bacillus subtilis functional genomics project which involved screening for sporulation genes, we identified an open reading frame, yaaT, whose disruptant exhibits a sporulation defect. Twenty-four hours after the initiation of sporulation, most cells of the yaaT mutant exhibited stage 0 of sporulation, indicating that the yaaT mutation blocks sporulation at an early stage. Furthermore, the mutation in yaaT led to a significant decrease in transcription from a promoter controlled by Spo0A, a key response regulator required for the initiation of sporulation. However, neither the level of transcription of spo0A, the activity of σH, which transcribes spo0A, nor the amount of Spo0A protein was severely affected by the mutation in yaaT. Bypassing the phosphorelay by introducing an spo0A mutation (sof-1) into the yaaT mutant suppressed the sporulation defect, suggesting that the yaaT mutation interferes with the phosphorelay process comprising Spo0F, Spo0B, and histidine kinases. We also observed that mutation of spo0E, which encodes the phosphatase that dephosphorylates Spo0A-P, suppressed the sporulation defect in the yaaT mutant. These results strongly suggest that yaaT plays a significant role in the transduction of signals to the phosphorelay for initiation of sporulation. Micrographs indicated that YaaT-green fluorescent protein localizes to the peripheral membrane, as well as to the septum, during sporulation.

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