Pseudomonas aeruginosa is a ubiquitous bacterium that survives in many environments, including as an acute and chronic pathogen in humans. Substantial evidence shows that P. aeruginosa behavior is affected by its motility, and appendages known as flagella and type IV pili (TFP) are known to confer such motility. The role these appendages play when not facilitating motility or attachment, however, is unclear. Here we discern a passive intercellular role of TFP during flagellar-mediated swarming of P. aeruginosa that does not require TFP extension or retraction. We studied swarming at the cellular level using a combination of laboratory experiments and computational simulations to explain the resultant patterns of cells imaged from in vitro swarms. Namely, we used a computational model to simulate swarming and to probe for individual cell behavior that cannot currently be otherwise measured. Our simulations showed that TFP of swarming P. aeruginosa should be distributed all over the cell and that TFP-TFP interactions between cells should be a dominant mechanism that promotes cell-cell interaction, limits lone cell movement, and slows swarm expansion. This predicted physical mechanism involving TFP was confirmed in vitro using pairwise mixtures of strains with and without TFP where cells without TFP separate from cells with TFP. While TFP slow swarm expansion, we show in vitro that TFP help alter collective motion to avoid toxic compounds such as the antibiotic carbenicillin. Thus, TFP physically affect P. aeruginosa swarming by actively promoting cell-cell association and directional collective motion within motile groups to aid their survival.

• Klíčová slova
• biofilms, collective motion, computational model, predictive simulations, self-organization,
• MeSH
• bakteriální adheze fyziologie   MeSH
• bakteriální fimbrie metabolismus   MeSH
• biofilmy růst a vývoj   MeSH
• biologické modely * MeSH
• červený fluorescenční protein MeSH
• flagella fyziologie   MeSH
• konfokální mikroskopie MeSH
• luminescentní proteiny MeSH
• mikrobiální interakce fyziologie   MeSH
• počítačová simulace MeSH
• pohyb fyziologie   MeSH
• Pseudomonas aeruginosa metabolismus   fyziologie   MeSH
• výpočetní biologie metody   MeSH
• zelené fluorescenční proteiny MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• luminescentní proteiny MeSH
• zelené fluorescenční proteiny MeSH

Porcine pleuropneumonia caused by Actinobacillus pleuropneumoniae accounts for serious economic losses in the pig farming industry worldwide. We examined here the immunogenicity and protective efficacy of the recombinant type IV fimbrial subunit protein ApfA as a single antigen vaccine against pleuropneumonia, or as a component of a multi-antigen preparation comprising five other recombinant antigens derived from key virulence factors of A. pleuropneumoniae (ApxIA, ApxIIA, ApxIIIA, ApxIVA and TbpB). Immunization of pigs with recombinant ApfA alone induced high levels of specific serum antibodies and provided partial protection against challenge with the heterologous A. pleuropneumoniae serotype 9 strain. This protection was higher than that engendered by vaccination with rApxIVA or rTbpB alone and similar to that observed after immunization with the tri-antigen combination of rApxIA, rApxIIA and rApxIIIA. In addition, rApfA improved the vaccination potential of the penta-antigen mixture of rApxIA, rApxIIA, rApxIIIA, rApxIVA and rTbpB proteins, where the hexa-antigen vaccine containing rApfA conferred a high level of protection on pigs against the disease. Moreover, when rApfA was used for vaccination alone or in combination with other antigens, such immunization reduced the number of pigs colonized with the challenge strain. These results indicate that ApfA could be a valuable component of an efficient subunit vaccine for the prevention of porcine pleuropneumonia.

• MeSH
• Actinobacillus pleuropneumoniae genetika   imunologie   patogenita   MeSH
• bakteriální adheziny genetika   imunologie   MeSH
• bakteriální fimbrie genetika   imunologie   MeSH
• bakteriální vakcíny genetika   imunologie   MeSH
• Escherichia coli genetika   MeSH
• exotoxiny genetika   metabolismus   MeSH
• faktory virulence genetika   imunologie   MeSH
• infekce bakteriemi rodu Actinobacillus imunologie   prevence a kontrola   veterinární   MeSH
• molekulární sekvence - údaje MeSH
• nemoci prasat imunologie   prevence a kontrola   MeSH
• pleuropneumonie imunologie   prevence a kontrola   veterinární   MeSH
• polymerázová řetězová reakce veterinární   MeSH
• prasata MeSH
• rekombinantní proteiny genetika   imunologie   MeSH
• sekvenční analýza DNA veterinární   MeSH
• vakcinace veterinární   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• randomizované kontrolované studie MeSH
• Názvy látek
• bakteriální adheziny MeSH
• bakteriální vakcíny MeSH
• exotoxiny MeSH
• faktory virulence MeSH
• rekombinantní proteiny MeSH

Type IV pilins are bacterial proteins that are small in size but have a broad range of functions, including motility, transformation competence and secretion. Although pilins vary in sequence, they possess a characteristic signal peptide that has to be removed by the prepilin peptidase PilD during pilin maturation. We generated a pilD (slr1120) null mutant of the cyanobacterium Synechocystis 6803 that accumulates an unprocessed form of the major pilin PilA1 (pPilA1) and its non-glycosylated derivative (NpPilA1). Notably, the pilD strain had aberrant membrane ultrastructure and did not grow photoautotrophically because the synthesis of Photosystem II subunits was abolished. However, other membrane components such as Photosystem I and ATP synthase were synthesized at levels comparable to the control strain. Proliferation of the pilD strain was rescued by elimination of the pilA1 gene, demonstrating that PilA1 prepilin inhibited the synthesis of Photosystem II. Furthermore, NpPilA1 co-immunoprecipitated with the SecY translocase and the YidC insertase, and both of these essential translocon components were degraded in the mutant. We propose that unprocessed prepilins inactivate an identical pool of translocons that function in the synthesis of both pilins and the core subunits of Photosystem II.

• MeSH
• bakteriální fimbrie metabolismus   MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• endopeptidasy genetika   metabolismus   MeSH
• fotosystém II - proteinový komplex metabolismus   MeSH
• glykosylace MeSH
• mutace MeSH
• proteiny fimbrií metabolismus   MeSH
• regulace genové exprese u bakterií MeSH
• Synechocystis genetika   růst a vývoj   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální proteiny MeSH
• endopeptidasy MeSH
• fotosystém II - proteinový komplex MeSH
• prepilin peptidase protein, Bacteria MeSH Prohlížeč
• proteiny fimbrií MeSH

Cyanobacteria synthesize type IV pili, which are known to be essential for motility, adhesion and natural competence. They consist of long flexible fibers that are primarily composed of the major pilin PilA1 in Synechocystis sp. PCC 6803. In addition, Synechocystis encodes less abundant pilin-like proteins, which are known as minor pilins. In this study, we show that the minor pilin PilA5 is essential for natural transformation but is dispensable for motility and flocculation. In contrast, a set of minor pilins encoded by the pilA9-slr2019 transcriptional unit are necessary for motility but are dispensable for natural transformation. Neither pilA5-pilA6 nor pilA9-slr2019 are essential for pilus assembly as mutant strains showed type IV pili on the cell surface. Three further gene products with similarity to PilX-like minor pilins have a function in flocculation of Synechocystis. The results of our study indicate that different minor pilins facilitate distinct pilus functions. Further, our microarray analysis demonstrated that the transcription levels of the minor pilin genes change in response to surface contact. A total of 122 genes were determined to have altered transcription between planktonic and surface growth, including several plasmid genes which are involved exopolysaccharide synthesis and the formation of bloom-like aggregates.

• Klíčová slova
• cyanobacteria, minor pilin, natural competence, surface acclimation, type IV pili,
• MeSH
• bakteriální fimbrie fyziologie   MeSH
• bakteriální proteiny fyziologie   MeSH
• fyziologie bakterií * MeSH
• mikročipová analýza MeSH
• proteiny fimbrií fyziologie   MeSH
• regulace genové exprese u bakterií MeSH
• sekvence aminokyselin MeSH
• sekvenční delece MeSH
• stanovení celkové genové exprese MeSH
• Synechocystis fyziologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální proteiny MeSH
• proteiny fimbrií MeSH

Bacteriophages are the most abundant biological entities on Earth, but our understanding of many aspects of their lifecycles is still incomplete. Here, we have structurally analysed the infection cycle of the siphophage Casadabanvirus JBD30. Using its baseplate, JBD30 attaches to Pseudomonas aeruginosa via the bacterial type IV pilus, whose subsequent retraction brings the phage to the bacterial cell surface. Cryo-electron microscopy structures of the baseplate-pilus complex show that the tripod of baseplate receptor-binding proteins attaches to the outer bacterial membrane. The tripod and baseplate then open to release three copies of the tape-measure protein, an event that is followed by DNA ejection. JBD30 major capsid proteins assemble into procapsids, which expand by 7% in diameter upon filling with phage dsDNA. The DNA-filled heads are finally joined with 180-nm-long tails, which bend easily because flexible loops mediate contacts between the successive discs of major tail proteins. It is likely that the structural features and replication mechanisms described here are conserved among siphophages that utilize the type IV pili for initial cell attachment.

• Klíčová slova
• Pseudomonas aeruginosa, Cryo-EM, Phage, Pili, Structure,
• MeSH
• bakteriální fimbrie metabolismus   ultrastruktura   virologie   MeSH
• DNA virů metabolismus   genetika   MeSH
• elektronová kryomikroskopie * MeSH
• fágy pseudomonád * ultrastruktura   genetika   metabolismus   fyziologie   MeSH
• Pseudomonas aeruginosa * virologie   metabolismus   MeSH
• replikace viru * MeSH
• Siphoviridae genetika   ultrastruktura   fyziologie   metabolismus   MeSH
• virové plášťové proteiny metabolismus   chemie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA virů MeSH
• virové plášťové proteiny MeSH

Type IV pili are bacterial surface-exposed filaments that are built up by small monomers called pilin proteins. Pilins are synthesized as longer precursors (prepilins), the N-terminal signal peptide of which must be removed by the processing protease PilD. A mutant of the cyanobacterium Synechocystis sp. PCC 6803 lacking the PilD protease is not capable of photoautotrophic growth because of the impaired function of Sec translocons. Here, we isolated phototrophic suppressor strains of the original ΔpilD mutant and, by sequencing their genomes, identified secondary mutations in the SigF sigma factor, the γ subunit of RNA polymerase, the signal peptide of major pilin PilA1, and in the pilA1-pilA2 intergenic region. Characterization of suppressor strains suggests that, rather than the total prepilin level in the cell, the presence of non-glycosylated PilA1 prepilin is specifically harmful. We propose that the restricted lateral mobility of the non-glycosylated PilA1 prepilin causes its accumulation in the translocon-rich membrane domains, which attenuates the synthesis of membrane proteins.

• Klíčová slova
• PilD peptidase, Synechocystis, Type IV pili, photosystem II, suppressor mutations,
• Publikační typ
• časopisecké články MeSH

FTH_0069 is a previously uncharacterized strongly immunoreactive protein that has been proposed to be a novel virulence factor in Francisella tularensis. Here, the glycan structure modifying two C-terminal peptides of FTH_0069 was identified utilizing high resolution, high mass accuracy mass spectrometry, combined with in-source CID tandem MS experiments. The glycan observed at m/z 1156 was determined to be a hexasaccharide, consisting of two hexoses, three N-acetylhexosamines, and an unknown monosaccharide containing a phosphate group. The monosaccharide sequence of the glycan is tentatively proposed as X-P-HexNAc-HexNAc-Hex-Hex-HexNAc, where X denotes the unknown monosaccharide. The glycan is identical to that of DsbA glycoprotein, as well as to one of the multiple glycan structures modifying the type IV pilin PilA, suggesting a common biosynthetic pathway for the protein modification. Here, we demonstrate that the glycosylation of FTH_0069, DsbA, and PilA was affected in an isogenic mutant with a disrupted wbtDEF gene cluster encoding O-antigen synthesis and in a mutant with a deleted pglA gene encoding pilin oligosaccharyltransferase PglA. Based on our findings, we propose that PglA is involved in both pilin and general F. tularensis protein glycosylation, and we further suggest an inter-relationship between the O-antigen and the glycan synthesis in the early steps in their biosynthetic pathways.

• MeSH
• faktory virulence chemie   genetika   metabolismus   MeSH
• Francisella tularensis genetika   metabolismus   patogenita   MeSH
• glykosylace MeSH
• molekulární sekvence - údaje MeSH
• multigenová rodina MeSH
• mutace MeSH
• O-antigeny chemie   genetika   metabolismus   MeSH
• proteiny fimbrií chemie   genetika   metabolismus   MeSH
• sacharidové sekvence MeSH
• sekvence aminokyselin MeSH
• tandemová hmotnostní spektrometrie MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• faktory virulence MeSH
• O-antigeny MeSH
• PilA protein, Francisella tularensis MeSH Prohlížeč
• proteiny fimbrií MeSH

In cyanobacteria that perform oxygenic photosynthesis, alternative sigma factors can play critical roles in environmental acclimation at the transcriptional initiation step. Here, we found in Synechococcus elongatus PCC 7942 that transcription of the pilA1 gene, encoding the type IV pilin, is dependent on one of the group 3 sigma factors, SigF1. We analyzed the promoter sequence determinants and proposed herein that the -10 and -35 boxes upstream of the transcriptional start site are critical for transcription. Interestingly, while the pilA1 promoter is activated by illumination, RNA polymerase containing SigF1 is already located on the promoter region under dark conditions, prior to illumination. This strongly suggests that promoter activation by light follows the recruitment of RNA polymerase during transcriptional initiation.

• Klíčová slova
• RNA polymerase, SigF1, cyanobacteria, pilA1 promoter, sigma factor, transcription,
• MeSH
• bakteriální proteiny * genetika   metabolismus   MeSH
• genetická transkripce MeSH
• promotorové oblasti (genetika) * MeSH
• proteiny fimbrií genetika   metabolismus   MeSH
• regulace genové exprese u bakterií * MeSH
• sigma faktor * genetika   metabolismus   MeSH
• světlo * MeSH
• Synechococcus * genetika   metabolismus   účinky záření   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny * MeSH
• proteiny fimbrií MeSH
• sigma faktor * MeSH

One motile, Gram-negative, non-spore-forming and rod-shaped symbiotic bacterium, strain UCH-936T, was isolated from Heterorhabditis atacamensis nematodes. Results of biochemical, physiological, molecular and genomic analyses suggest that it represents a new species, which we propose to name Photorhabdus antumapuensis sp. nov. Digital DNA-DNA hybridization shows that strain UCH-936T is more closely related to Photorhabdus kleinii DSM 23513T, but shares solely 50.5 % similarity, which is below the 70% cut-off value that delimits species boundaries in bacteria. Phylogenetic reconstructions using whole-genome sequences show that strain UCH-936T forms a unique clade, suggesting its novel and distinct taxonomic status again. Similarly, comparative genomic analyses shows that the virulence factor flagella-related gene fleR, the type IV pili-related gene pilL and the vibriobactin-related gene vibE are present in the genome of strain UCH-936T but absent in the genomes of its closest relatives. Biochemically and physiologically, UCH-936T differs also from all closely related Photorhabdus species. Therefore, Photorhabdus antumapuensis sp. nov. is proposed as a new species with the type strain UCH-936T (CCCT 21.06T=CCM 9188T=CCOS 1991T).

• Klíčová slova
• Heterorhabditis, Photorhabdus, entomopathogenic bacteria, symbiotic bacteria, taxonomy,
• MeSH
• DNA bakterií genetika   MeSH
• faktory virulence MeSH
• fylogeneze MeSH
• hlístice * MeSH
• mastné kyseliny chemie   MeSH
• Photorhabdus * genetika   MeSH
• Rhabditoidea * mikrobiologie   MeSH
• RNA ribozomální 16S genetika   MeSH
• sekvenční analýza DNA MeSH
• techniky typizace bakterií MeSH
• zastoupení bazí MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA bakterií MeSH
• faktory virulence MeSH
• mastné kyseliny MeSH
• RNA ribozomální 16S MeSH

The role of fibronectin (Fn) and its natural receptors alpha5beta1 integrins in the interaction of P. aeruginosa with A549 epithelial cells was compared in the clinical isolate ER97314 and the reference PAK strain. Both strains expressed functional type IV pili, as shown by the results of the twitching motility assay. The ER97314 strain was highly adherent to immobilized Fn (640 000+/-20 000 CFU per well) while the PAK strain adhered less efficiently (70 000+/-10 000 CFU per well). Both strains adhered to A549 cells (33 400+/-1200 and 1200+/-100 CFU per well, for PAK and ER97314, respectively), only the PAK strain being significantly internalized (9430+/-2020 CFU per well). Cytochalasin D and genistein significantly decreased bacterial adherence of the 2 strains and caused also a significant decrease in PAK internalization. This inhibitory activity was not related to changes in the expression of alpha5beta1 integrins. Antibodies to Fn and alpha5beta1 integrins inhibited the adherence of the ER97314 strain but had no significant effect on PAK interaction with human cells. These findings suggest that only some P. aeruginosa strains can target Fn and their natural receptors alpha5beta1 integrins for adherence to A549 cells.

• MeSH
• bakteriální adheze * MeSH
• buněčné linie MeSH
• epitelové buňky mikrobiologie   MeSH
• fibronektiny metabolismus   MeSH
• integrin alfa5beta1 metabolismus   MeSH
• lidé MeSH
• Pseudomonas aeruginosa patogenita   fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fibronektiny MeSH
• integrin alfa5beta1 MeSH