Binding of exogenous DNA to marine sediments and the effect of DNA/sediment binding on natural transformation ofPseudomonas stutzeristrain ZoBell in sediment columns

ABSTRACTThe increasing frequency of bacteria showing antimicrobial resistance (AMR) raises the menace of entering into a postantibiotic era. Horizontal gene transfer (HGT) is one of the prime reasons for AMR acquisition.Acinetobacter baumanniiis a nosocomial pathogen with outstanding abilities to survive in the hospital environment and to acquire resistance determinants. Its capacity to incorporate exogenous DNA is a major source of AMR genes; however, few studies have addressed this subject. The transformation machinery as well as the factors that induce natural competence inA. baumanniiare unknown. In this study, we demonstrate that naturally competent strain A118 increases its natural transformation frequency upon the addition of Ca2+or albumin. We show thatcomEAandpilQare involved in this process since their expression levels are increased upon the addition of these compounds. An unspecific protein, like casein, does not reproduce this effect, showing that albumin's effect is specific. Our work describes the first specific inducers of natural competence inA. baumannii. Overall, our results suggest that the main protein in blood enhances HGT inA. baumannii, contributing to the increase of AMR in this threatening human pathogen.

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CryoEM structure of the type IVa pilus secretin required for natural competence in Vibrio cholerae

Nature Communications ◽

10.1038/s41467-020-18866-y ◽

2020 ◽

Vol 11 (1) ◽

Author(s):

Sara J. Weaver ◽

Davi R. Ortega ◽

Matthew H. Sazinsky ◽

Triana N. Dalia ◽

Ankur B. Dalia ◽

...

Keyword(s):

Antibiotic Resistance ◽

Gene Transfer ◽

Horizontal Gene Transfer ◽

Vibrio Cholerae ◽

Natural Transformation ◽

Domain Architecture ◽

Genetic Material ◽

Surface Binding ◽

Exogenous Dna ◽

Insight Into

Abstract Natural transformation is the process by which bacteria take up genetic material from their environment and integrate it into their genome by homologous recombination. It represents one mode of horizontal gene transfer and contributes to the spread of traits like antibiotic resistance. In Vibrio cholerae, a type IVa pilus (T4aP) is thought to facilitate natural transformation by extending from the cell surface, binding to exogenous DNA, and retracting to thread this DNA through the outer membrane secretin, PilQ. Here, we use a functional tagged allele of VcPilQ purified from native V. cholerae cells to determine the cryoEM structure of the VcPilQ secretin in amphipol to ~2.7 Å. We use bioinformatics to examine the domain architecture and gene neighborhood of T4aP secretins in Proteobacteria in comparison with VcPilQ. This structure highlights differences in the architecture of the T4aP secretin from the type II and type III secretion system secretins. Based on our cryoEM structure, we design a series of mutants to reversibly regulate VcPilQ gate dynamics. These experiments support the idea of VcPilQ as a potential druggable target and provide insight into the channel that DNA likely traverses to promote the spread of antibiotic resistance via horizontal gene transfer by natural transformation.

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Silencing of natural transformation by an RNA chaperone and a multitarget small RNA

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1601626113 ◽

2016 ◽

Vol 113 (31) ◽

pp. 8813-8818 ◽

Cited By ~ 60

Author(s):

Laetitia Attaiech ◽

Aïda Boughammoura ◽

Céline Brochier-Armanet ◽

Omran Allatif ◽

Flora Peillard-Fiorente ◽

...

Keyword(s):

Legionella Pneumophila ◽

Natural Transformation ◽

Bacterial Species ◽

Physiological State ◽

Uptake System ◽

Dna Uptake ◽

Exogenous Dna ◽

Major Mechanism ◽

Transcriptional Gene Regulation ◽

High Level

A highly conserved DNA uptake system allows many bacteria to actively import and integrate exogenous DNA. This process, called natural transformation, represents a major mechanism of horizontal gene transfer (HGT) involved in the acquisition of virulence and antibiotic resistance determinants. Despite evidence of HGT and the high level of conservation of the genes coding the DNA uptake system, most bacterial species appear non-transformable under laboratory conditions. In naturally transformable species, the DNA uptake system is only expressed when bacteria enter a physiological state called competence, which develops under specific conditions. Here, we investigated the mechanism that controls expression of the DNA uptake system in the human pathogenLegionella pneumophila. We found that a repressor of this system displays a conserved ProQ/FinO domain and interacts with a newly characterizedtrans-acting sRNA, RocR. Together, they target mRNAs of the genes coding the DNA uptake system to control natural transformation. This RNA-based silencing represents a previously unknown regulatory means to control this major mechanism of HGT. Importantly, these findings also show that chromosome-encoded ProQ/FinO domain-containing proteins can assisttrans-acting sRNAs and that this class of RNA chaperones could play key roles in post-transcriptional gene regulation throughout bacterial species.

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CryoEM structure of the Vibrio cholerae Type IV competence pilus secretin PilQ

10.1101/2020.03.03.975896 ◽

2020 ◽

Cited By ~ 1

Author(s):

Sara J. Weaver ◽

Matthew H. Sazinsky ◽

Triana N. Dalia ◽

Ankur B. Dalia ◽

Grant J. Jensen

Keyword(s):

Antibiotic Resistance ◽

Gene Transfer ◽

Horizontal Gene Transfer ◽

Vibrio Cholerae ◽

Natural Transformation ◽

Structural Information ◽

Genetic Material ◽

Surface Binding ◽

Exogenous Dna ◽

Type Iv

AbstractNatural transformation is the process by which bacteria take up genetic material from their environment and integrate it into their genome by homologous recombination. It represents one mode of horizontal gene transfer and contributes to the spread of traits like antibiotic resistance. In Vibrio cholerae, the Type IV competence pilus is thought to facilitate natural transformation by extending from the cell surface, binding to exogenous DNA, and retracting to thread this DNA through the outer membrane secretin, PilQ. A lack of structural information has hindered our understanding of this process, however. Here, we solved the first ever high-resolution structure of a Type IV competence pilus secretin. A functional tagged allele of VcPilQ purified from native V. cholerae cells was used to determine the cryoEM structure of the PilQ secretin in amphipol to ∼2.7 Å. This structure highlights for the first time key differences in the architecture of the Type IV competence pilus secretin from the Type II and Type III Secretin System secretins. Based on our cryoEM structure, we designed a series of mutants to interrogate the mechanism of PilQ. These experiments provide insight into the channel that DNA likely traverses to promote the spread of antibiotic resistance via horizontal gene transfer by natural transformation. We prove that it is possible to reduce pilus biogenesis and natural transformation by sealing the gate, suggesting VcPilQ as a new drug target.

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Competence for Natural Transformation Is Common among Clinical Strains of Resistant Acinetobacter spp.

Microorganisms ◽

10.3390/microorganisms7020030 ◽

2019 ◽

Vol 7 (2) ◽

pp. 30 ◽

Cited By ~ 11

Author(s):

Sara Domingues ◽

Natasha Rosário ◽

Ângela Cândido ◽

Daniela Neto ◽

Kaare Nielsen ◽

...

Keyword(s):

Gene Transfer ◽

Natural Transformation ◽

Clinical Isolates ◽

Bacterial Populations ◽

Acinetobacter Species ◽

Exogenous Dna ◽

Time Period ◽

Resistance Patterns ◽

Important Trait ◽

Acinetobacter Spp

Horizontal gene transfer events provide the basis for extensive dissemination of antimicrobial resistance traits between bacterial populations. Conjugation is considered to be the most frequent mechanism behind new resistance acquisitions in clinical pathogens but does not fully explain the resistance patterns seen in some bacterial genera. Gene transfer by natural transformation has been described for numerous clinical isolates, including some Acinetobacter species. The main aim of this study was to determine to what extent clinical, resistant Acinetobacter spp. isolates, express competence for natural transformation. Twenty-two clinical Acinetobacter spp. isolates collected over a 16-year time period, from five different geographical separated and/or distinct Portuguese Hospitals were tested for natural transformability. Fourteen isolates, including 11 A. baumannii, 2 A. nosocomialis and 1 Acinetobacter sp., were identified as competent on semisolid media facilitating surface-motility. Competent Acinetobacter isolates were found in all the hospitals tested. Furthermore, osmolarity was shown to influence the uptake of exogenous DNA by competent A. baumannii A118. Our study demonstrates that natural competence is common among clinical isolates of Acinetobacter spp., and hence likely an important trait for resistance acquisition.

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Noncanonical Cell-to-Cell DNA Transfer in Thermus spp. Is Insensitive to Argonaute-Mediated Interference

Journal of Bacteriology ◽

10.1128/jb.02113-14 ◽

2014 ◽

Vol 197 (1) ◽

pp. 138-146 ◽

Cited By ~ 14

Author(s):

Alba Blesa ◽

Carolina Elvira César ◽

Beate Averhoff ◽

José Berenguer

Keyword(s):

Natural Transformation ◽

Thermus Thermophilus ◽

Rapid Evolution ◽

Extreme Thermophile ◽

Bacterial Populations ◽

Exogenous Dna ◽

Content Type ◽

Naked Dna ◽

Transfer Mechanisms ◽

Dna Acquisition

Horizontal gene transfer drives the rapid evolution of bacterial populations. Classical processes that promote the lateral flow of genetic information are conserved throughout the prokaryotic world. However, some species have nonconserved transfer mechanisms that are not well known. This is the case for the ancient extreme thermophileThermus thermophilus. In this work, we show thatT. thermophilusstrains are capable of exchanging large DNA fragments by a novel mechanism that requires cell-to-cell contacts and employs components of the natural transformation machinery. This process facilitates the bidirectional transfer of virtually any DNA locus but favors by 10-fold loci found in the megaplasmid over those in the chromosome. In contrast to naked DNA acquisition by transformation, the system does not activate the recently described DNA-DNA interference mechanism mediated by the prokaryotic Argonaute protein, thus allowing the organism to distinguish between DNA transferred from a mate and exogenous DNA acquired from unknown hosts. This Argonaute-mediated discrimination may be tentatively viewed as a strategy for safe sharing of potentially “useful” traits by the components of a given population ofThermusspp. without increasing the genome sizes of its individuals.

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CrfP, a fratricide protein, contributes to natural transformation in Streptococcus suis

Veterinary Research ◽

10.1186/s13567-021-00917-x ◽

2021 ◽

Vol 52 (1) ◽

Author(s):

Yinchu Zhu ◽

Jiale Ma ◽

Yue Zhang ◽

Xiaojun Zhong ◽

Qiankun Bai ◽

...

Keyword(s):

Natural Transformation ◽

Potential Effect ◽

Transformation Process ◽

Transformation Rate ◽

Bacterial Cells ◽

Exogenous Dna ◽

Src Homology 3 ◽

Src Homology ◽

Natural Genetic Transformation

AbstractStreptococcussuis (S.suis) is an important zoonotic pathogen that causes septicaemia, meningitis and streptococcal toxic shock-like syndrome in its host, and recent studies have shown that S.suis could be competent for natural genetic transformation. Transformation is an important mechanism for the horizontal transfer of DNA, but some elements that affect the transformation process need to be further explored. Upon entering the competent state, Streptococcus species stimulate the transcription of competence-related genes that are responsible for exogenous DNA binding, uptake and processing. In this study, we performed conserved promoter motif and qRT-PCR analyses and identified CrfP as a novel murein hydrolase that is widespread in S.suis and stimulated with a peptide pheromone in the competent state through a process controlled by ComX. A bioinformatics analysis revealed that CrfP consists of a CHAP hydrolase domain and two bacterial Src homology 3-binding (SH3b) domains. Further characterization showed that CrfP could be exported to extracellular bacterial cells and lytic S.suis strains of different serotypes, and this finding was verified by TEM and a turbidity assay. To investigate the potential effect of CrfP in vivo, a gene-deletion mutant (ΔcrfP) was constructed. Instead of stopping the natural transformation process, the inactivation of CrfP clearly reduced the effective transformation rate. Overall, these findings provide evidence showing that CrfP is important for S.suis serovar 2 competence.

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Identification of neisserial DNA binding components

Microbiology ◽

10.1099/mic.0.022640-0 ◽

2009 ◽

Vol 155 (3) ◽

pp. 852-862 ◽

Cited By ~ 18

Author(s):

Emma Lång ◽

Kristine Haugen ◽

Burkhard Fleckenstein ◽

Håvard Homberset ◽

Stephan A. Frye ◽

...

Keyword(s):

Dna Binding ◽

Natural Transformation ◽

Solid Phase ◽

Binding Activity ◽

Dna Uptake ◽

Inner Membrane ◽

Exogenous Dna ◽

Dna Binding Activity ◽

Pilus Biogenesis ◽

Null Mutants

Neisseria meningitidis, a causative agent of meningitis and septicaemia, expresses type IV pili, a feature correlating with the uptake of exogenous DNA from the environment by natural transformation. The outer membrane complex PilQ, through which pili are extruded and retracted, has previously been shown to bind DNA in its pore region. In order to further elucidate how DNA is transported across the membranes, we searched for DNA binding proteins within the meningococcal inner membrane. Inner membrane fractions from a panel of neisserial strains were subjected to a solid-phase overlay assay with DNA substrates, and MS was subsequently employed to identify proteins that bind DNA. A number of DNA binding components were detected, including the pilus biogenesis component PilG, the competence protein ComL, and the cell division ATP-binding protein FtsE, as well as two hypothetical proteins. The DNA binding activity of these components was not dependent on the presence of the neisserial DNA uptake sequence. Null mutants, corresponding to each of the proteins identified, were constructed to assess their phenotypes. Only mutants defective in pilus biogenesis were non-competent and non-piliated. The DNA binding activity of the pilus biogenesis components PilQ and PilG and the phenotypes of their respective null mutants suggest that these proteins are directly involved as players in natural transformation, and not only indirectly, through pilus biogenesis.

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Vibrio campbellii DS40M4 is a tractable model strain that diverges from the canonical quorum-sensing regulatory circuit in vibrios

10.1101/2020.03.31.019307 ◽

2020 ◽

Author(s):

Chelsea A. Simpson ◽

Blake D. Petersen ◽

Logan J. Geyman ◽

Aimee H. Lee ◽

Michael P. Manzella ◽

...

Keyword(s):

Quorum Sensing ◽

Small Rnas ◽

Natural Transformation ◽

Response Regulator ◽

Exogenous Dna ◽

Sensing System ◽

Quorum Sensing System ◽

Vibrio Campbellii ◽

Type Strains ◽

Model Strain

AbstractVibrio campbellii BB120 (previously designated as Vibrio harveyi) is a fundamental model strain for studying population density-based cell-to-cell communication, known as quorum sensing. In V. campbellii BB120, sensing of autoinducers at high cell densities activates the expression of the master transcriptional regulator, LuxR, which controls the expression of genes involved in group behaviors. The environmental isolate Vibrio campbellii DS40M4 was recently shown to be capable of natural transformation, a process by which bacteria take up exogenous DNA and incorporate it into their genome via homologous recombination. In contrast, BB120 is not naturally transformable. Here, we compare additional phenotypes between these two V. campbellii strains. DS40M4 has a faster growth rate and stronger type VI secretion-mediated cell killing, whereas BB120 forms more robust biofilms and is bioluminescent. To explore the function of DS40M4-encoded homologs of the BB120 quorum-sensing system, we exploited the power of natural transformation to rapidly generate >30 mutant strains. Our results show that DS40M4 has a similar quorum-sensing circuit to BB120 but with three distinct differences: 1) DS40M4 lacks the canonical HAI-1 autoinducer LuxM synthase but has an active LuxN receptor, 2) the quorum regulatory small RNAs (Qrrs) are not solely regulated by autoinducer signaling through the response regulator LuxO, and 3) the DS40M4 LuxR regulon is <100 genes, which is relatively small compared to the >400 genes regulated in BB120. This work illustrates that DS40M4 is a tractable and relevant model strain for studying quorum-sensing phenotypes in Vibrio campbellii.ImportanceWild isolates of bacterial type strains can yield important information about traits that vary within species. Here, we compare the recently sequenced isolate of Vibrio campbellii DS40M4 to the canonical lab type strain BB120 and examine several phenotypes that define this species, including quorum sensing, bioluminescence, and biofilm formation. Importantly, DS40M4 is naturally transformable with exogenous DNA, which allows for the rapid generation of mutants in a laboratory setting. By exploiting natural transformation, we genetically dissected the functions of BB120 quorum-sensing system homologs in the DS40M4 strain, including two-component signaling systems, transcriptional regulators, and small RNAs. Our results show important distinctions between the quorum-sensing circuits of these two strains that underscore the need to examine wild isolates alongside type strains.

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