RESUMO
Adherence to both cellular and abiotic surfaces is a crucial step in the interaction of bacterial pathogens and commensals with their hosts. Bacterial surface structures known as fimbriae or pili play a fundamental role in the early colonization stages by providing specificity or tropism. Among the various fimbrial families, the chaperone-usher family has been extensively studied due to its ubiquity, diversity, and abundance. This family is named after the components that facilitate their biogenesis. Type 1 fimbria and P pilus, two chaperone-usher fimbriae associated with urinary tract infections, have been thoroughly investigated and serve as prototypes that have laid the foundations for understanding the biogenesis of this fimbrial family. Additionally, the study of the mechanisms regulating their expression has also been a subject of great interest, revealing that the regulation of the expression of the genes encoding these structures is a complex and diverse process, involving both common global regulators and those specific to each operon.
Assuntos
Proteínas de Fímbrias , Fímbrias Bacterianas , Regulação Bacteriana da Expressão Gênica , Chaperonas Moleculares , Fímbrias Bacterianas/metabolismo , Fímbrias Bacterianas/genética , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/genética , Proteínas de Fímbrias/genética , Proteínas de Fímbrias/metabolismo , Aderência Bacteriana , ÓperonRESUMO
Background. Scant information is available regarding fimbrillins within the genus Porphyromonas, with the notable exception of those belonging to Porphyromonas gingivalis, which have been extensively researched for several years. Besides fim and mfa, a third P. gingivalis adhesin called filament-forming protein 1 (Ffp1) has recently been described and seems to be pivotal for outer membrane vesicle (OMV) production. Objective. We aimed to investigate the distribution and diversity of type V fimbrillin, particularly Ffp1, in the genus Porphyromonas. Methods. A bioinformatics phylogenomic analysis was conducted using all accessible Porphyromonas genomes to generate a domain search for fimbriae, using hidden Markov model profiles. Results. Ffp1 was identified as the sole fimbrillin present in all analysed genomes. After manual verification (i.e. biocuration) of both structural and functional annotations and 3D modelling, this protein was determined to be a type V fimbrillin, with a closer structural resemblance to a Bacteroides ovatus fimbrillin than to FimA or Mfa1 from P. gingivalis. Conclusion. It appears that Ffp1 is an ancestral fimbria, transmitted through vertical inheritance and present across all Porphyromonas species. Additional investigations are necessary to elucidate the biogenesis of Ffp1 fimbriae and their potential role in OMV production and niche adaptation.
RESUMO
In Escherichia coli, the disaccharide trehalose can be metabolized as a carbon source or be accumulated as an osmoprotectant under osmotic stress. In hypertonic environments, E. coli accumulates trehalose in the cell by synthesis from glucose mediated by the cytosolic enzymes OtsA and OtsB. Trehalose in the periplasm can be hydrolyzed into glucose by the periplasmic trehalase TreA. We have previously shown that a treA mutant of extraintestinal E. coli strain BEN2908 displayed increased resistance to osmotic stress by 0.6 M urea, and reduced production of type 1 fimbriae, reduced invasion of avian fibroblasts, and decreased bladder colonization in a murine model of urinary tract infection. Since loss of TreA likely results in higher periplasmic trehalose concentrations, we wondered if deletion of otsA and otsB genes, which would lead to decreased internal trehalose concentrations, would reduce resistance to stress by 0.6 M urea and promote type 1 fimbriae production. The BEN2908ΔotsBA mutant was sensitive to osmotic stress by urea, but displayed an even more pronounced reduction in production of type 1 fimbriae, with the consequent reduction in adhesion/invasion of avian fibroblasts and reduced bladder colonization in the murine urinary tract. The BEN2908ΔtreAotsBA mutant also showed a reduction in production of type 1 fimbriae, but in contrast to the ΔotsBA mutant, resisted better than the wild type in the presence of urea. We hypothesize that, in BEN2908, resistance to stress by urea would depend on the levels of periplasmic trehalose, but type 1 fimbriae production would be influenced by the levels of cytosolic trehalose.
Assuntos
Fímbrias Bacterianas , Osmorregulação , Trealose , Bexiga Urinária , Infecções Urinárias , Animais , Trealose/metabolismo , Camundongos , Bexiga Urinária/microbiologia , Fímbrias Bacterianas/metabolismo , Fímbrias Bacterianas/genética , Infecções Urinárias/microbiologia , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/genética , Modelos Animais de Doenças , Feminino , Pressão Osmótica , Escherichia coli Extraintestinal Patogênica/metabolismo , Escherichia coli Extraintestinal Patogênica/genética , Ureia/metabolismo , Trealase/metabolismo , Trealase/genética , Deleção de Genes , Glucose/metabolismoRESUMO
Background and Aims: The pathogenicity of Escherichia coli is determined by the presence of genes that mediate virulence factors such as adherence capacity and toxin production. This research aimed to identify the adhesion factors and antibiotic resistance capacity of E. coli strains associated with diarrhea in piglets in Colombia. Materials and Methods: Presumptive E. coli strains were isolated from the rectal swabs of piglets in swine farms between 4 and 40 days of age with evidence of diarrhea. Presumptive E. coli strains were tested for antibiotic resistance. The hemolytic capacity of presumptive E. coli strains was measured and molecularly identified. Strains confirmed as hemolytic E. coli was evaluated for the presence of five adhesion factors (F4, F5, F6, F18, and F41) and resistance to 11 antibiotics. Results: Fifty-two putative E. coli strains were isolated, six of which showed a hemolytic capacity. The hemolytic strains were molecularly identified as E. coli. Adhesive fimbriae were found in five of six ß-hemolytic E. coli isolates. Combinations of the adhesion factors F6-F18 and F6-F41 were linked to antibiotic resistance capacity. Conclusion: The phenomenon of E. coli strains resistant to multiple antibiotics on pig farms represents a constant risk factor for public health and pig production.
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Introduction: Gallibacterium anatis causes gallibacteriosis in birds. These bacteria produce biofilms and secrete several fimbrial appendages as tools to cause disease in animals. G. anatis strains contain up to three types of fimbriae. Complete genome sequencing is the strategy currently used to determine variations in the gene content of G. anatis, although today only the completely circularized genome of G. anatis UMN179 is available. Methods: The appearance of growth of various strains of G. anatis in liquid culture medium was studied. Biofilm production and how the amount of biofilm was affected by DNase, Proteinase K, and Pronase E enzymes were analyzed. Fimbrial gene expression was performed by protein analysis and qRT-PCR. In an avian model, the pathogenesis generated by the strains G. anatis ESV200 and 12656-12 was investigated. Using bioinformatic tools, the complete genome of G. anatis ESV200 was comparatively studied to search for virulence factors that would help explain the pathogenic behavior of this strain. Results and Discussion: G. anatis ESV200 strain differs from the 12656-12 strain because it produces a biofilm at 20%. G. anatis ESV200 strain express fimbrial genes and produces biofilm but with a different structure than that observed for strain 12656-12. ESV200 and 12656-12 strains are pathogenic for chickens, although the latter is the most virulent. Here, we show that the complete genome of the ESV200 strain is similar to that of the UNM179 strain. However, these strains have evolved with many structural rearrangements; the most striking chromosomal arrangement is a Maverick-like element present in the ESV200 strain.
RESUMO
Enteroaggregative Escherichia coli (EAEC) is an important cause of diarrhea in children and adults worldwide. This pathotype is phenotypically characterized by the aggregative-adherence (AA) pattern in HEp-2 cells and genetically associated to the presence of the aatA gene. EAEC pathogenesis relies in different virulence factors. At least, three types of adhesins have been specifically associated with EAEC strains: the five variants of the aggregative adherence fimbriae (AAF), the aggregative forming pilus (AFP) and more recently, a fibrilar adhesin named CS22. Our study aimed to evaluate the presence of AAF, AFP and CS22-related genes among 110 EAEC strains collected from feces of children with diarrhea. The presence of aggR (EAEC virulence regulator) and genes related to AAFs (aggA, aafA, agg3A, agg4A, agg5A and agg3/4C), AFP (afpA1 and afpR) and CS22 (cseA) was detected by PCR, and the adherence patterns were evaluated on HeLa cells. aggR-positive strains comprised 83.6% of the collection; among them, 80.4% carried at least one AAF-related gene and presented the AA pattern. aggA was the most frequent AAF-related gene (28.4% of aggR+ strains). cseA was detected among aggR+ (16.3%) and aggR- strains (22.2%); non-adherent strains or strains presenting AA pattern were observed in both groups. afpR and afpA1 were exclusively detected among aggR- strains (77.8%), most of which (71.4%) also presented AA pattern. Our results indicate that AAF- and AFP-related genes may contribute to identify EAEC strains, while the presence of cseA and its importance as an EAEC virulence factor and genotypic marker needs to be further evaluated.
Assuntos
Adesinas Bacterianas , Escherichia coli , alfa-Fetoproteínas , Criança , Humanos , Adesinas Bacterianas/genética , Aderência Bacteriana/genética , Biomarcadores , Diarreia/microbiologia , Escherichia coli/genética , Células HeLa , Fatores de Virulência/genéticaRESUMO
Pil-fimbriae is a type IV pili member, which is a remarkably versatile component with a wide variety of functions, including motility, attachment to different surfaces, electrical conductance, DNA acquisition, and secretion of a broad range of structurally distinct protein substrates. Despite the previous functional characterization of Pil, more studies are required to understand the regulation of Pil expression and production, since the exact mechanisms involved in these steps are still unknown. Therefore it is extremely important to have a protein with the correct secondary and tertiary structure that will enable an accurate characterization and a specific antisera generation. For this reason, the aim of this work was to generate potential tools for further investigations to comprehend the mechanisms involved in Pil regulation and its role in pathogenic E. coli infections with the obtaining of a precise native-like recombinant PilS and the corresponding antisera. The pilS gene was successfully cloned into an expression vector, and recombinant PilS (rPilS) was efficiently solubilized and purified by metal affinity chromatography. Protein characterization analyses indicated that rPilS presented native-like secondary and tertiary structures after the refolding process. The generated anti-rPilS sera efficiently recognized recombinant and native proteins from atypical enteropathogenic E. coli strains.
RESUMO
Enteroaggregative Escherichia coli (EAEC) is an important cause of diarrhea in children and adults worldwide. This pathotype is phenotypically characterized by the aggregative-adherence (AA) pattern in HEp-2 cells and genetically associated to the presence of the aatA gene. EAEC pathogenesis relies in different virulence factors. At least, three types of adhesins have been specifically associated with EAEC strains: the five variants of the aggregative adherence fimbriae (AAF), the aggregative forming pilus (AFP) and more recently, a fibrilar adhesin named CS22. Our study aimed to evaluate the presence of AAF, AFP and CS22-related genes among 110 EAEC strains collected from feces of children with diarrhea. The presence of aggR (EAEC virulence regulator) and genes related to AAFs (aggA, aafA, agg3A, agg4A, agg5A and agg3/4C), AFP (afpA1 and afpR) and CS22 (cseA) was detected by PCR, and the adherence patterns were evaluated on HeLa cells. aggR-positive strains comprised 83.6% of the collection; among them, 80.4% carried at least one AAF-related gene and presented the AA pattern. aggA was the most frequent AAF-related gene (28.4% of aggR+ strains). cseA was detected among aggR+ (16.3%) and aggR- strains (22.2%); non-adherent strains or strains presenting AA pattern were observed in both groups. afpR and afpA1 were exclusively detected among aggR- strains (77.8%), most of which (71.4%) also presented AA pattern. Our results indicate that AAF- and AFP-related genes may contribute to identify EAEC strains, while the presence of cseA and its importance as an EAEC virulence factor and genotypic marker needs to be further evaluated.
RESUMO
Pil-fimbriae is a type IV pili member, which is a remarkably versatile component with a wide variety of functions, including motility, attachment to different surfaces, electrical conductance, DNA acquisition, and secretion of a broad range of structurally distinct protein substrates. Despite the previous functional characterization of Pil, more studies are required to understand the regulation of Pil expression and production, since the exact mechanisms involved in these steps are still unknown. Therefore it is extremely important to have a protein with the correct secondary and tertiary structure that will enable an accurate characterization and a specific antisera generation. For this reason, the aim of this work was to generate potential tools for further investigations to comprehend the mechanisms involved in Pil regulation and its role in pathogenic E. coli infections with the obtaining of a precise native-like recombinant PilS and the corresponding antisera. The pilS gene was successfully cloned into an expression vector, and recombinant PilS (rPilS) was efficiently solubilized and purified by metal affinity chromatography. Protein characterization analyses indicated that rPilS presented native-like secondary and tertiary structures after the refolding process. The generated anti-rPilS sera efficiently recognized recombinant and native proteins from atypical enteropathogenic E. coli strains.