scholarly journals Tad pilus-mediated twitching motility is essential for DNA uptake and survival of Liberibacters

PLoS ONE ◽  
2021 ◽  
Vol 16 (10) ◽  
pp. e0258583
Author(s):  
Lulu Cai ◽  
Mukesh Jain ◽  
Marta Sena-Vélez ◽  
Kathryn M. Jones ◽  
Laura A. Fleites ◽  
...  
Keyword(s):  
Natural Transformation ◽  
Gene Repertoire ◽  
Dna Uptake ◽  
Pathogenic Species ◽  
Twitching Motility ◽  
Inner Membrane ◽  
Nucleotide Biosynthesis ◽  
Genes Encoding ◽  
Conserved Gene ◽  
First Time

Axenically cultured Liberibacter crescens (Lcr) is a closely related surrogate for uncultured plant pathogenic species of the genus Liberibacter, including ‘Candidatus L. asiaticus’ (CLas) and ‘Ca. L. solanacearum’ (CLso). All Liberibacters encode a completely conserved gene repertoire for both flagella and Tad (Tight Adherence) pili and all are missing genes critical for nucleotide biosynthesis. Both flagellar swimming and Tad pilus-mediated twitching motility in Lcr were demonstrated for the first time. A role for Tad pili in the uptake of extracellular dsDNA for food in Liberibacters was suspected because both twitching and DNA uptake are impossible without repetitive pilus extension and retraction, and no genes encoding other pilus assemblages or mechanisms for DNA uptake were predicted to be even partially present in any of the 35 fully sequenced Liberibacter genomes. Insertional mutations of the Lcr Tad pilus genes cpaA, cpaB, cpaE, cpaF and tadC all displayed such severely reduced growth and viability that none could be complemented. A mutation affecting cpaF (motor ATPase) was further characterized and the strain displayed concomitant loss of twitching, viability and reduced periplasmic uptake of extracellular dsDNA. Mutations of comEC, encoding the inner membrane competence channel, had no effect on either motility or growth but completely abolished natural transformation in Lcr. The comEC mutation was restored by complementation using comEC from Lcr but not from CLas strain psy62 or CLso strain RS100, indicating that unlike Lcr, these pathogens were not naturally competent for transformation. This report provides the first evidence that the Liberibacter Tad pili are dynamic and essential for both motility and DNA uptake, thus extending their role beyond surface adherence.

scholarly journals Requirement of Novel Competence Genes pilT andpilU of Pseudomonas stutzeri for Natural Transformation and Suppression of pilT Deficiency by a Hexahistidine Tag on the Type IV Pilus Protein PilAI

2001 ◽  
Vol 183 (16) ◽  
pp. 4694-4701 ◽  
Author(s):  
Stefan Graupner ◽  
Nicole Weger ◽  
Monika Sohni ◽  
Wilfried Wackernagel
Keyword(s):  
Amino Acid ◽  
Natural Transformation ◽  
Pseudomonas Stutzeri ◽  
Amino Acid Identity ◽  
Binding Motif ◽  
Dna Uptake ◽  
Twitching Motility ◽  
Type Iv Pilus ◽  
Type Iv ◽  
Insertional Inactivation

ABSTRACT The ubiquitous species Pseudomonas stutzeri has type IV pili, and these are essential for the natural transformation of the cells. An absolute transformation-deficient mutant obtained after transposon mutagenesis had an insertion in a gene which was termedpilT. The deduced amino acid sequence has identity with PilT of Pseudomonas aeruginosa (94%), Neisseria gonorrhoeae (67%), and other gram-negative species and it contains a nucleotide-binding motif. The mutant was hyperpiliated but defective for further pilus-associated properties, such as twitching motility and plating of pilus-specific phage PO4. [3H]thymidine-labeled DNA was bound by the mutant but not taken up. Downstream of pilT a gene, termedpilU, coding for a putative protein with 88% amino acid identity with PilU of P. aeruginosa was identified. Insertional inactivation did not affect piliation, twitching motility, or PO4 infection but reduced transformation to about 10%. The defect was fully complemented by PilU of nontransformable P. aeruginosa. When thepilAI gene (coding for the type IV pilus prepilin) was manipulated to code for a protein in which the six C-terminal amino acids were replaced by six histidine residues and then expressed from a plasmid, it gave a nonpiliated and twitching motility-defective phenotype in pilAI::Gmr cells but allowed transformability. Moreover, the mutant allele suppressed the absolute transformation deficiency caused by the pilT mutation. Considering the hypothesized role of pilT + in pilus retraction and the presumed requirement of retraction for DNA uptake, it is proposed that the pilT-independent transformation is promoted by PilA mutant protein either as single molecules or as minimal pilin assembly structures in the periplasm which may resemble depolymerized pili and that these cause the outer membrane pores to open for DNA entry.

scholarly journals Type IV Pilus Biogenesis, Twitching Motility, and DNA Uptake in Thermus thermophilus: Discrete Roles of Antagonistic ATPases PilF, PilT1, and PilT2

2013 ◽  
Vol 80 (2) ◽  
pp. 644-652 ◽  
Author(s):  
Ralf Salzer ◽  
Friederike Joos ◽  
Beate Averhoff
Keyword(s):  
Natural Transformation ◽  
Thermophilic Bacteria ◽  
Bacterial Species ◽  
Thermus Thermophilus ◽  
Dna Uptake ◽  
Twitching Motility ◽  
Type Iv Pilus ◽  
Content Type ◽  
Ecological Diversification ◽  
Type Iv

ABSTRACTNatural transformation has a large impact on lateral gene flow and has contributed significantly to the ecological diversification and adaptation of bacterial species.Thermus thermophilusHB27 has emerged as the leading model organism for studies of DNA transporters in thermophilic bacteria. Recently, we identified a zinc-binding polymerization nucleoside triphosphatase (NTPase), PilF, which is essential for the transport of DNA through the outer membrane. Here, we present genetic evidence that PilF is also essential for the biogenesis of pili. One of the most challenging questions was whetherT. thermophilushas any depolymerization NTPase acting as a counterplayer of PilF. We identified two depolymerization NTPases, PilT1 (TTC1621) and PilT2 (TTC1415), both of which are required for type IV pilus (T4P)-mediated twitching motility and adhesion but dispensable for natural transformation. This suggests that T4P dynamics are not required for natural transformation. The latter finding is consistent with our suggestion that inT. thermophilus, T4P and natural transformation are linked but distinct systems.

scholarly journals Identification of neisserial DNA binding components

Microbiology ◽  
2009 ◽  
Vol 155 (3) ◽  
pp. 852-862 ◽  
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.

scholarly journals Pilin-Like Proteins in the Extremely Thermophilic Bacterium Thermus thermophilus HB27: Implication in Competence for Natural Transformation and Links to Type IV Pilus Biogenesis

2003 ◽  
Vol 69 (7) ◽  
pp. 3695-3700 ◽  
Author(s):  
Alexandra Friedrich ◽  
Judit Rumszauer ◽  
Anke Henne ◽  
Beate Averhoff
Keyword(s):  
Natural Transformation ◽  
Thermus Thermophilus ◽  
Thermophilic Bacterium ◽  
Type Iv Pili ◽  
Dna Uptake ◽  
Significant Similarity ◽  
High Frequencies ◽  
Type Iv ◽  
Genes Encoding ◽  
Pilus Biogenesis

ABSTRACT The extreme thermophile Thermus thermophilus HB27 exhibits high frequencies of natural transformation. Although we recently reported identification of the first competence genes in Thermus, the molecular basis of DNA uptake is unknown. A pilus-like structure is assumed to be involved. Twelve genes encoding prepilin-like proteins were identified in three loci in the genome of T. thermophilus. Mutational analyses, described in this paper, revealed that one locus, which contains four genes that encode prepilin-like proteins (pilA1 to pilA4), is essential for natural transformation. Additionally, comZ, a new competence gene with no similarity to known genes, was identified. Analysis of the piliation phenotype revealed wild-type piliation of a pilA1-pilA3Δkat mutant and a comZ mutant, whereas a pilA4 mutant was found to be completely devoid of pilus structures. These findings, together with the significant similarity of PilA4 to prepilins, led to the conclusion that the T. thermophilus pilus structures are type IV pili. Furthermore, the loss of the transformation and piliation phenotype in the pilA4 mutant suggests that type IV pili are implicated in natural transformation of T. thermophilus HB27.

scholarly journals Twitching and Swimming Motility Play a Role in Ralstonia solanacearum Pathogenicity

mSphere ◽  
2020 ◽  
Vol 5 (2) ◽  
Author(s):  
Jordi Corral ◽  
Pau Sebastià ◽  
Núria S. Coll ◽  
Jordi Barbé ◽  
Jesús Aranda ◽  
...  
Keyword(s):  
Plant Pathogen ◽  
Ralstonia Solanacearum ◽  
Biofilm Formation ◽  
Natural Transformation ◽  
Plant Colonization ◽  
Twitching Motility ◽  
Tomato Plants ◽  
Content Type ◽  
First Time ◽  
Root Attachment

ABSTRACT Ralstonia solanacearum is a bacterial plant pathogen causing important economic losses worldwide. In addition to the polar flagella responsible for swimming motility, this pathogen produces type IV pili (TFP) that govern twitching motility, a flagellum-independent movement on solid surfaces. The implication of chemotaxis in plant colonization, through the control flagellar rotation by the proteins CheW and CheA, has been previously reported in R. solanacearum. In this work, we have identified in this bacterium homologues of the Pseudomonas aeruginosa pilI and chpA genes, suggested to play roles in TFP-associated motility analogous to those played by the cheW and cheA genes, respectively. We demonstrate that R. solanacearum strains with a deletion of the pilI or the chpA coding region show normal swimming and chemotaxis but altered biofilm formation and reduced twitching motility, transformation efficiency, and root attachment. Furthermore, these mutants displayed wild-type growth in planta and impaired virulence on tomato plants after soil-drench inoculations but not when directly applied to the xylem. Comparison with deletion mutants for pilA and fliC—encoding the major pilin and flagellin subunits, respectively—showed that both twitching and swimming are required for plant colonization and full virulence. This work proves for the first time the functionality of a pilus-mediated pathway encoded by pil-chp genes in R. solanacearum, demonstrating that pilI and chpA genes are bona fide motility regulators controlling twitching motility and its three related phenotypes: virulence, natural transformation, and biofilm formation. IMPORTANCE Twitching and swimming are two bacterial movements governed by pili and flagella. The present work identifies for the first time in the Gram-negative plant pathogen Ralstonia solanacearum a pilus-mediated chemotaxis pathway analogous to that governing flagellum-mediated chemotaxis. We show that regulatory genes in this pathway control all of the phenotypes related to pili, including twitching motility, natural transformation, and biofilm formation, and are also directly implicated in virulence, mainly during the first steps of the plant infection. Our results show that pili have a higher impact than flagella on the interaction of R. solanacearum with tomato plants and reveal new types of cross-talk between the swimming and twitching motility phenotypes: enhanced swimming in bacteria lacking pili and a role for the flagellum in root attachment.

scholarly journals Uncovering dynamic evolution in the plastid genome of seven Ligusticum species provides insights into species discrimination and phylogenetic implications

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Can Yuan ◽  
Xiufen Sha ◽  
Miao Xiong ◽  
Wenjuan Zhong ◽  
Yu Wei ◽  
...  
Keyword(s):  
Plastid Genome ◽  
Species Discrimination ◽  
Chloroplast Genomes ◽  
Genomic Arrangement ◽  
Phylogenetic Implications ◽  
Solid Foundation ◽  
Conserved Gene ◽  
Fine Resolution ◽  
Genome Scale ◽  
First Time

AbstractLigusticum L., one of the largest members in Apiaceae, encompasses medicinally important plants, the taxonomic statuses of which have been proved to be difficult to resolve. In the current study, the complete chloroplast genomes of seven crucial plants of the best-known herbs in Ligusticum were presented. The seven genomes ranged from 148,275 to 148,564 bp in length with a highly conserved gene content, gene order and genomic arrangement. A shared dramatic decrease in genome size resulted from a lineage-specific inverted repeat (IR) contraction, which could potentially be a promising diagnostic character for taxonomic investigation of Ligusticum, was discovered, without affecting the synonymous rate. Although a higher variability was uncovered in hotspot divergence regions that were unevenly distributed across the chloroplast genome, a concatenated strategy for rapid species identification was proposed because separate fragments inadequately provided variation for fine resolution. Phylogenetic inference using plastid genome-scale data produced a concordant topology receiving a robust support value, which revealed that L. chuanxiong had a closer relationship with L. jeholense than L. sinense, and L. sinense cv. Fuxiong had a closer relationship to L. sinense than L. chuanxiong, for the first time. Our results not only furnish concrete evidence for clarifying Ligusticum taxonomy but also provide a solid foundation for further pharmaphylogenetic investigation.

scholarly journals The Assessment of IL-21 and IL-22 at the mRNA Level in Tumor Tissue and Protein Concentration in Serum and Peritoneal Fluid in Patients with Ovarian Cancer

2021 ◽  
Vol 10 (14) ◽  
pp. 3058
Author(s):  
Aleksandra Mielczarek-Palacz ◽  
Celina Kruszniewska-Rajs ◽  
Marta Smycz-Kubańska ◽  
Jarosław Strzelczyk ◽  
Wojciech Szanecki ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Peritoneal Fluid ◽  
Tumor Tissue ◽  
Mrna Level ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
First Time ◽  
Ovarian Serous Cancer

The aim of the analysis was for the first time to assess the expression of genes encoding IL-21 and IL-22 at the mRNA level in ovarian tumor specimens and the concentration of these parameters in serum and peritoneal fluid in patients with ovarian serous cancer. The levels of IL-21 and IL-22 transcripts were evaluated with the use of the real-time RT-qPCR. Enzyme-linked immunosorbent assay (ELISA) was used to determine the concentration of proteins. Quantitative analysis of IL-21 gene mRNA in the tumor tissue showed the highest activity in the G1 degree of histopathological differentiation and was higher in G1 compared to the control group. The concentration of IL-21 and IL-22 in the serum and in the peritoneal fluid of women with ovarian cancer varied depending on the degree of histopathological differentiation of the cancer and showed statistical variability compared to controls. The conducted studies have shown that the local and systemic changes in the immune system involving IL-21 and IL-22 indicate the participation of these parameters in the pathogenesis of ovarian cancer, and modulation in the IL-21/IL-22 system may prove useful in the development of new diagnostic and therapeutic strategies used in patients, which require further research.

scholarly journals Designing P. aeruginosa synthetic phages with reduced genomes

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Diana P. Pires ◽  
Rodrigo Monteiro ◽  
Dalila Mil-Homens ◽  
Arsénio Fialho ◽  
Timothy K. Lu ◽  
...  
Keyword(s):  
Galleria Mellonella ◽  
Antibacterial Properties ◽  
Genetically Engineered ◽  
In Vivo Studies ◽  
Infection Model ◽  
Promising Therapeutic Approach ◽  
Genes Encoding ◽  
First Time

AbstractIn the era where antibiotic resistance is considered one of the major worldwide concerns, bacteriophages have emerged as a promising therapeutic approach to deal with this problem. Genetically engineered bacteriophages can enable enhanced anti-bacterial functionalities, but require cloning additional genes into the phage genomes, which might be challenging due to the DNA encapsulation capacity of a phage. To tackle this issue, we designed and assembled for the first time synthetic phages with smaller genomes by knocking out up to 48% of the genes encoding hypothetical proteins from the genome of the newly isolated Pseudomonas aeruginosa phage vB_PaeP_PE3. The antibacterial efficacy of the wild-type and the synthetic phages was assessed in vitro as well as in vivo using a Galleria mellonella infection model. Overall, both in vitro and in vivo studies revealed that the knock-outs made in phage genome do not impair the antibacterial properties of the synthetic phages, indicating that this could be a good strategy to clear space from phage genomes in order to enable the introduction of other genes of interest that can potentiate the future treatment of P. aeruginosa infections.

scholarly journals A Deadly Cargo: Gene Repertoire of Cytotoxic Effector Proteins in the Camelidae

Genes ◽  
2021 ◽  
Vol 12 (2) ◽  
pp. 304
Author(s):  
Ján Futas ◽  
Jan Oppelt ◽  
Pamela Anna Burger ◽  
Petr Horin
Keyword(s):  
Cytotoxic T Cells ◽  
Effector Proteins ◽  
Target Cells ◽  
Gene Repertoire ◽  
Interspecific Differences ◽  
Genes Encoding ◽  
Bactrian Camels ◽  
Pore Forming Proteins ◽  
Sequence Similarities ◽  
Cytotoxic Effector

Cytotoxic T cells and natural killer cells can kill target cells based on their expression and release of perforin, granulysin, and granzymes. Genes encoding these molecules have been only poorly annotated in camelids. Based on bioinformatic analyses of genomic resources, sequences corresponding to perforin, granulysin, and granzymes were identified in genomes of camelids and related ungulate species, and annotation of the corresponding genes was performed. A phylogenetic tree was constructed to study evolutionary relationships between the species analyzed. Re-sequencing of all genes in a panel of 10 dromedaries and 10 domestic Bactrian camels allowed analyzing their individual genetic polymorphisms. The data showed that all extant Old World camelids possess functional genes for two pore-forming proteins (PRF1, GNLY) and six granzymes (GZMA, GZMB, GZMH, GZMK, GZMM, and GZMO). All these genes were represented as single copies in the genome except the GZMH gene exhibiting interspecific differences in the number of loci. High protein sequence similarities with other camelid and ungulate species were observed for GZMK and GZMM. The protein variability in dromedaries and Bactrian camels was rather low, except for GNLY and chymotrypsin-like granzymes (GZMB, GZMH).

scholarly journals Prevalence of Eimeria spp. in calves from dairy farms in northern Paraná state, Brazil

2018 ◽  
Vol 27 (1) ◽  
pp. 118-122
Author(s):  
Sérgio Tosi Cardim ◽  
Mércia Seixas ◽  
Victor Bittencourt Dutra Tabacow ◽  
Alessandra Taroda ◽  
Priscilla Gomes Carneiro ◽  
...  
Keyword(s):  
Species Identification ◽  
Dairy Farms ◽  
Northern Region ◽  
The State ◽  
Pathogenic Species ◽  
Eimeria Spp ◽  
High Prevalence ◽  
The World ◽  
Cattle Herds ◽  
First Time

Abstract Bovine coccidiosis is a disease of major importance in cattle herds across the world. The disorder mainly affects young calves, and E. bovis and E. zuernii are considered the most pathogenic species of the genus, however, E. alabamensis have been described in grazing calves. In this study, the prevalence of Eimeria spp. was evaluated in calves on dairy farms in the northern region of the state of Paraná, Brazil. Four hundred calves on 44 dairy farms were tested for the presence of coccidian oocysts. The positives were re-examined and the oocysts were morphometrically analyzed for species identification. All the farms were contaminated and 205 animals (51.25%) presented Eimeria spp. oocysts. Among these, 146 animals (71.22%) were co-infected by two or more species of coccidia. Ten species of Eimeria were identified: E. bovis (in 30.25% of the positive samples), E. alabamensis (26.75%), E. zuernii (22.00%), E. ellipsoidalis (18.50%), E. auburnensis (13.75%), E. canadensis (8.00%), E. cylindrica (7.25%), E. subspherica (5.00%), E. bukidnonensis (3.00%) and E. brasiliensis (0.75%). This study demonstrates the high prevalence of Eimeria spp. in the northern region of Paraná, Brazil, and detection for the first time in our region the pathogenic species E. alabamensis.

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