RESUMEN
miRNAs are major regulators of eukaryotic gene expression and host immunity, and play an important role in the inflammation-mediated pathways in periodontal disease (PD) pathogenesis. Expanding our previous observation with the global miRNA profiling using partial human mouth microbes, and lack of in vivo studies involving oral spirochete Treponema denticola-induced miRNAs, this study was designed to delineate the global miRNA expression kinetics during progression of periodontitis in mice infected with T. denticola by using NanoString nCounter® miRNA panels. All of the T. denticola-infected male and female mice at 8 and 16 weeks demonstrated bacterial colonization (100%) on the gingival surface, and an increase in alveolar bone resorption (p < 0.0001). A total of 70 miRNAs with at least 1.0-fold differential expression/regulation (DE) (26 upregulated and 44 downregulated) were identified. nCounter miRNA expression profiling identified 13 upregulated miRNAs (e.g., miR-133a, miR-378) and 25 downregulated miRNAs (e.g., miR-375, miR-34b-5p) in T. denticola-infected mouse mandibles during 8 weeks of infection, whereas 13 upregulated miRNAs (e.g., miR-486, miR-126-5p) and 19 downregulated miRNAs (miR-2135, miR-142-3p) were observed during 16 weeks of infection. One miRNA (miR-126-5p) showed significant difference between 8 and 16 weeks of infection. Interestingly, miR-126-5p has been presented as a potential biomarker in patients with periodontitis and coronary artery disease. Among the upregulated miRNAs, miR-486, miR-126-3p, miR-126-5p, miR-378a-3p, miR-22-3p, miR-151a-3p, miR-423-5p, and miR-221 were reported in human gingival plaques and saliva samples from periodontitis and with diabetes. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed various functional pathways of DE miRNAs, such as bacterial invasion of epithelial cells, Ras signaling, Fc gamma R-mediated phagocytosis, osteoclast differentiation, adherens signaling, and ubiquitin mediated proteolysis. This is the first study of DE miRNAs in mouse mandibles at different time-points of T. denticola infection; the combination of three specific miRNAs, miR-486, miR-126-3p, and miR-126-5p, may serve as an invasive biomarker of T. denticola in PD. These miRNAs may have a significant role in PD pathogenesis, and this research establishes a link between miRNA, periodontitis, and systemic diseases.
Asunto(s)
Enfermedades Transmisibles , MicroARNs , Enfermedades Periodontales , Periodontitis , Humanos , Masculino , Femenino , Animales , Ratones , MicroARNs/genética , MicroARNs/metabolismo , Treponema denticola/genética , Spirochaetales/genética , Treponema/genética , Treponema/metabolismo , Cinética , Perfilación de la Expresión Génica , Periodontitis/genética , Enfermedades Periodontales/genética , BiomarcadoresRESUMEN
Treponema denticola, a keystone pathogen in periodontitis, is a model organism for studying Treponema physiology and host-microbe interactions. Its major surface protein Msp forms an oligomeric outer membrane complex that binds fibronectin, has cytotoxic pore-forming activity, and disrupts several intracellular processes in host cells. T. denticola msp is an ortholog of the Treponema pallidum tprA to -K gene family that includes tprK, whose remarkable in vivo hypervariability is proposed to contribute to T. pallidum immune evasion. We recently identified the primary Msp surface-exposed epitope and proposed a model of the Msp protein as a ß-barrel protein similar to Gram-negative bacterial porins. Here, we report fine-scale Msp mutagenesis demonstrating that both the N and C termini as well as the centrally located Msp surface epitope are required for native Msp oligomer expression. Removal of as few as three C-terminal amino acids abrogated Msp detection on the T. denticola cell surface, and deletion of four residues resulted in complete loss of detectable Msp. Substitution of a FLAG tag for either residues 6 to 13 of mature Msp or an 8-residue portion of the central Msp surface epitope resulted in expression of full-length Msp but absence of the oligomer, suggesting roles for both domains in oligomer formation. Consistent with previously reported Msp N-glycosylation, proteinase K treatment of intact cells released a 25 kDa polypeptide containing the Msp surface epitope into culture supernatants. Molecular modeling of Msp using novel metagenome-derived multiple sequence alignment (MSA) algorithms supports the hypothesis that Msp is a large-diameter, trimeric outer membrane porin-like protein whose potential transport substrate remains to be identified. IMPORTANCE The Treponema denticola gene encoding its major surface protein (Msp) is an ortholog of the T. pallidum tprA to -K gene family that includes tprK, whose remarkable in vivo hypervariability is proposed to contribute to T. pallidum immune evasion. Using a combined strategy of fine-scale mutagenesis and advanced predictive molecular modeling, we characterized the Msp protein and present a high-confidence model of its structure as an oligomer embedded in the outer membrane. This work adds to knowledge of Msp-like proteins in oral treponemes and may contribute to understanding the evolutionary and potential functional relationships between T. denticola Msp and the orthologous T. pallidum Tpr proteins.
Asunto(s)
Fibronectinas , Treponema denticola , Aminoácidos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Endopeptidasa K/metabolismo , Epítopos , Fibronectinas/metabolismo , Péptidos/metabolismo , Porinas/metabolismo , Treponema/química , Treponema/genética , Treponema/metabolismo , Treponema denticola/genéticaRESUMEN
The outer membrane proteins (OMPs) of Treponema pallidum subsp. pallidum (T. pallidum), the etiological agent of the sexually transmitted disease syphilis, have long been a hot research topic. Despite many hurdles to studying the pathogen, especially the inability to manipulate T. pallidum in vitro genetically, considerable progress has been made in elucidating the structure, pathogenesis and functions of T. pallidum OMPs. In this review, we integrate this information to garner fresh insights into the role of OMPs in the diagnosis, pathogenicity and vaccine development of T. pallidum. Collectively, the essential scientific discussions herein should provide a framework for understanding the current status and prospects of T. pallidum OMPs.
Asunto(s)
Sífilis , Treponema pallidum , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas de la Membrana Bacteriana Externa/metabolismo , Humanos , Sífilis/diagnóstico , Treponema/metabolismo , Treponema pallidum/genéticaRESUMEN
The prokaryotic chemotaxis system is arguably the best-understood signaling pathway in biology. In all previously described species, chemoreceptors organize into a hexagonal (P6 symmetry) extended array. Here, we report an alternative symmetry (P2) of the chemotaxis apparatus that emerges from a strict linear organization of the histidine kinase CheA in Treponema denticola cells, which possesses arrays with the highest native curvature investigated thus far. Using cryo-ET, we reveal that Td chemoreceptor arrays assume an unusual arrangement of the supra-molecular protein assembly that has likely evolved to accommodate the high membrane curvature. The arrays have several atypical features, such as an extended dimerization domain of CheA and a variant CheW-CheR-like fusion protein that is critical for maintaining an ordered chemosensory apparatus. Furthermore, the previously characterized Td oxygen sensor ODP influences CheA ordering. These results suggest a greater diversity of the chemotaxis signaling system than previously thought.
Asunto(s)
Membrana Celular/metabolismo , Células Quimiorreceptoras/citología , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Membrana Celular/ultraestructura , Células Quimiorreceptoras/metabolismo , Quimiotaxis , Secuencia Conservada , Escherichia coli/citología , Eliminación de Gen , Histidina Quinasa/metabolismo , Dominios Proteicos , Homología de Secuencia de Aminoácido , Treponema/metabolismoRESUMEN
Functional coupling and comparative genomics analysis have been applied to study functional associations of orthologs of enterococcal cAD1 sex pheromone (P13268) known to be responsible for biofilm formation, conjugative plasmid transfer and spreading of bacterial antibiotics resistance. cAD1 peptide pheromone is released from the membrane lipoprotein with the peptide precursor encoded by a gene cad (tr|C2JQE7). Our analysis of genomic neighbourhood of cad and motifs of the encoded polypeptide and its orthologs suggests a close functional association between cAD1 and ApbE protein (Q82Z24), a FMN insertion and trafficking facilitator. The cad and apbE orthologs were coupled in the genomes and ApbE-specific motifs for FMN covalent attachment were identified in cad-encoded protein sequence and its orthologs. These findings suggest a potential role of FMN-based reductase function of the cAD1 lipoprotein precursor in its processing and release of the active sex pheromone peptide. They may lead to a new approach in prevention of antibiotic resistance spread via targeting sex pheromone processing chaperones or by suppression of the FMN availability and covalent binding. This methods can be also applied to a controlled evolution of bacterial pathogenicity in microbial fuel cells, as the findings suggest the crosstalk between bacterial pathogenicity and bacterial electro-activity.
Asunto(s)
Proteínas Bacterianas , Enterobacteriaceae , Chaperonas Moleculares , Oligopéptidos , Procesamiento Proteico-Postraduccional/fisiología , Treponema , Secuencias de Aminoácidos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Enterobacteriaceae/genética , Enterobacteriaceae/metabolismo , Lipoproteínas/genética , Lipoproteínas/metabolismo , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Oligopéptidos/genética , Oligopéptidos/metabolismo , Treponema/genética , Treponema/metabolismoRESUMEN
The cecum plays an important role in the feed fermentation of ruminants. However, information is very limited regarding the cecal microbiota and their methane production. In the present study, the cecal content from twelve local Chinese goats, fed with either a hay diet (0% grain) or a high-grain diet (71.5% grain), were used to investigate the bacterial and archaeal community and their methanogenic potential. Microbial community analysis was determined using high-throughput sequencing of 16S rRNA genes and real-time PCR, and the methanogenesis potential was assessed by in vitro fermentation with ground corn or hay as substrates. Compared with the hay group, the high-grain diet significantly increased the length and weight of the cecum, the proportions of starch and crude protein, the concentrations of volatile fatty acids and ammonia nitrogen, but decreased the pH values (P < 0.05). The high-grain diet significantly increased the abundances of bacteria and archaea (P < 0.05) and altered their community. For the bacterial community, the genera Bifidobacterium, Prevotella, and Treponema were significantly increased in the high-grain group (P < 0.05), while Akkermansia, Oscillospira, and Coprococcus were significantly decreased (P < 0.05). For the archaeal community, Methanosphaera stadtmanae was significantly increased in the high-grain group (P < 0.05), while Methanosphaera sp. ISO3-F5 was significantly decreased (P < 0.05). In the in vitro fermentation with grain as substrate, the cecal microorganisms from the high-grain group produced a significantly higher amount of methane and volatile fatty acids (P < 0.05), and produced significantly lower amount of lactate (P < 0.05). Conclusively, high-grain diet led to more fermentable substrates flowing into the hindgut of goats, resulting in an enhancement of microbial fermentation and methane production in the cecum.
Asunto(s)
Archaea/genética , Bacterias/genética , Ciego/microbiología , Grano Comestible , Animales , Archaea/clasificación , Archaea/metabolismo , Bacterias/clasificación , Bacterias/metabolismo , Bifidobacterium/clasificación , Bifidobacterium/genética , Bifidobacterium/metabolismo , Biología Computacional , Ácidos Grasos Volátiles/metabolismo , Cabras , Metano/metabolismo , Methanobacteriaceae/citología , Methanobacteriaceae/genética , Methanobacteriaceae/metabolismo , Prevotella/clasificación , Prevotella/genética , Prevotella/metabolismo , ARN Ribosómico 16S/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Treponema/clasificación , Treponema/genética , Treponema/metabolismoRESUMEN
Spirochetes are suspected to be linked to the genesis of neurological diseases, including neurosyphillis or neurodegeneration (ND). Impaired iron homeostasis has been implicated in loss of function in several enzymes requiring iron as a cofactor, formation of toxic oxidative species, inflammation and elevated production of beta-amyloid proteins. This review proposes to discuss the link that may exist between the involvement of Treponema spp. in the genesis or worsening of ND, and iron dyshomeostasis. Proteins secreted by Treponema can act directly on iron metabolism, with hemin binding ability (HbpA and HbpB) and iron reductase able to reduce the central ferric iron of hemin, iron-containing proteins (rubredoxin, neelaredoxin, desulfoferrodoxin metalloproteins, bacterioferritins etc). Treponema can also interact with cellular compounds, especially plasma proteins involved in iron metabolism, contributing to the virulence of the syphilis spirochetes (e.g. treponemal motility and survival). Fibronectin, transferrin and lactoferrin were also shown to be receptors for treponemal adherence to host cells and extracellular matrix. Association between Treponema and iron binding proteins results in iron accumulation and sequestration by Treponema from host macromolecules during systemic and mucosal infections.
Asunto(s)
Hierro/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Treponema/metabolismo , Infecciones por Treponema/metabolismo , Animales , Proteínas Bacterianas/metabolismo , Grupo Citocromo b/metabolismo , Ferritinas/metabolismo , Humanos , Enfermedades Neurodegenerativas/epidemiología , Enfermedades Neurodegenerativas/microbiología , Spirochaetales/aislamiento & purificación , Spirochaetales/metabolismo , Transferrina/metabolismo , Treponema/aislamiento & purificación , Infecciones por Treponema/epidemiologíaRESUMEN
The spirochete Treponema pallidum subsp. pallidum is the causative agent of syphilis, a chronic, sexually transmitted infection characterized by multiple symptomatic and asymptomatic stages. Although several other species in the genus are able to cause or contribute to disease, T. pallidum differs in that it is able to rapidly disseminate via the bloodstream to tissue sites distant from the site of initial infection. It is also the only Treponema species able to cross both the blood-brain and placental barriers. Previously, the T. pallidum proteins, Tp0750 and Tp0751 (also called pallilysin), were shown to degrade host proteins central to blood coagulation and basement membrane integrity, suggesting a role for these proteins in T. pallidum dissemination and tissue invasion. In the present study, we characterized Tp0750 and Tp0751 sequence variation in a diversity of pathogenic and nonpathogenic treponemes. We also determined the proteolytic potential of the orthologs from the less invasive species Treponema denticola and Treponema phagedenis. These analyses showed high levels of sequence similarity among Tp0750 orthologs from pathogenic species. For pallilysin, lower levels of sequence conservation were observed between this protein and orthologs from other treponemes, except for the ortholog from the highly invasive rabbit venereal syphilis-causing Treponema paraluiscuniculi. In vitro host component binding and degradation assays demonstrated that pallilysin and Tp0750 orthologs from the less invasive treponemes tested were not capable of binding or degrading host proteins. The results show that pallilysin and Tp0750 host protein binding and degradative capability is positively correlated with treponemal invasiveness.
Asunto(s)
Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Sífilis/metabolismo , Treponema pallidum/metabolismo , Secuencia de Aminoácidos , Animales , Proteínas Bacterianas/química , Secuencia Conservada , Humanos , Datos de Secuencia Molecular , Filogenia , Proteolisis , Conejos , Alineación de Secuencia , Especificidad de la Especie , Sífilis/microbiología , Treponema/clasificación , Treponema/genética , Treponema/metabolismo , Treponema/patogenicidad , Treponema pallidum/clasificación , Treponema pallidum/genética , Treponema pallidum/patogenicidad , VirulenciaRESUMEN
Pectin is a non-fiber carbohydrate (NFC) that exists in forages, but it is not clear how pectin exerts its effect on populations of either known microbial species or uncultured ruminal bacteria. PCR-denaturing gradient gel electrophoresis (PCR-DGGE) and real-time PCR analysis were used in the present study to investigate the effects of pectin on microbial communities in an in vitro rumen fermentation system. The fermentations were conducted using forage (corn stover or alfalfa), an NFC source (pectin or corn starch), or their combination as the substrates. Addition of pectin increased acetate (P < 0.05), whereas inclusion of starch increased butyrate production (P < 0.05). The pectate lyase activity was higher with alfalfa than with corn straw, or with pectin than with corn starch (P < 0.05), while the amylase activity was higher in corn starch-included treatments than the others (P < 0.05). The cluster analysis of the bacterial 16S rRNA gene showed that the DGGE banding patterns differed significantly between the treatments and led to the identification of three groups that were highly associated with the NFC sources. The specific bands associated with pectin-rich treatments were identified to be dominated by members of the Treponema genus. The growth of the Treponema genus was remarkably supported by the inclusion of pectin, highlighting their specific ability to degrade pectin. The results from the present study expand our knowledge of the microbial populations associated with pectin digestion, which may not only facilitate future research on utilization of pectin in feeds, but also improve our understanding of pectin digestion with respect to the rumen micro-ecosystem.
Asunto(s)
Bacterias/aislamiento & purificación , Microbiota , Pectinas/metabolismo , Rumen/microbiología , Treponema/aislamiento & purificación , Animales , Bacterias/clasificación , Bacterias/genética , Bacterias/metabolismo , Fermentación , Técnicas In Vitro , Filogenia , Rumen/metabolismo , Treponema/clasificación , Treponema/genética , Treponema/metabolismoRESUMEN
UNLABELLED: When prokaryotic cells acquire mutations, encounter translation-inhibiting substances, or experience adverse environmental conditions that limit their ability to synthesize proteins, transcription can become uncoupled from translation. Such uncoupling is known to suppress transcription of protein-encoding genes in bacteria. Here we show that the trace element selenium controls transcription of the gene for the selenocysteine-utilizing enzyme formate dehydrogenase (fdhFSec) through a translation-coupled mechanism in the termite gut symbiont Treponema primitia, a member of the bacterial phylum Spirochaetes. We also evaluated changes in genome-wide transcriptional patterns caused by selenium limitation and by generally uncoupling translation from transcription via antibiotic-mediated inhibition of protein synthesis. We observed that inhibiting protein synthesis in T. primitia influences transcriptional patterns in unexpected ways. In addition to suppressing transcription of certain genes, the expected consequence of inhibiting protein synthesis, we found numerous examples in which transcription of genes and operons is truncated far downstream from putative promoters, is unchanged, or is even stimulated overall. These results indicate that gene regulation in bacteria allows for specific post-initiation transcriptional responses during periods of limited protein synthesis, which may depend both on translational coupling and on unclassified intrinsic elements of protein-encoding genes. IMPORTANCE: A large body of literature demonstrates that the coupling of transcription and translation is a general and essential method by which bacteria regulate gene expression levels. However, the potential role of noncanonical amino acids in regulating transcriptional output via translational control remains, for the most part, undefined. Furthermore, the genome-wide transcriptional state in response to translational decoupling is not well quantified. The results presented here suggest that the noncanonical amino acid selenocysteine is able to tune transcription of an important metabolic gene via translational coupling. Furthermore, a genome-wide analysis reveals that transcriptional decoupling produces a wide-ranging effect and that this effect is not uniform. These results exemplify how growth conditions that impact translational processivity can rapidly feed back on transcriptional productivity of prespecified groups of genes, providing bacteria with an efficient response to environmental changes.
Asunto(s)
Biosíntesis de Proteínas/efectos de los fármacos , Selenio/metabolismo , Transcripción Genética/efectos de los fármacos , Treponema/efectos de los fármacos , Treponema/metabolismo , Animales , Formiato Deshidrogenasas/metabolismo , Tracto Gastrointestinal/microbiología , Perfilación de la Expresión Génica , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Isópteros/microbiología , Treponema/genéticaRESUMEN
A new method for simultaneous coke oven gas (COG) biomethanation and in situ biogas upgrading in anaerobic reactor was developed in this study. The simulated coke oven gas (SCOG) (92% H2 and 8% CO) was injected directly into the anaerobic reactor treating sewage sludge through hollow fiber membrane (HFM). With pH control at 8.0, the added H2 and CO were fully consumed and no negative effects on the anaerobic degradation of sewage sludge were observed. The maximum CH4 content in the biogas was 99%. The addition of SCOG resulted in enrichment and dominance of homoacetogenetic genus Treponema and hydrogenotrophic genus Methanoculleus in the liquid, which indicated that H2 were converted to methane by both direct (hydrogenotrophic methanogenesis) and indirect (homoacetogenesis+aceticlastic methanogenesis) pathways in the liquid. However, the aceticlasitic genus Methanosaeta was dominant for archaea in the biofilm on the HFM, which indicated indirect (homoacetogenesis+aceticlastic methanogenesis) H2 conversion pathway on the biofilm.
Asunto(s)
Contaminantes Atmosféricos/química , Biocombustibles , Reactores Biológicos , Coque , Anaerobiosis , Biopelículas , Monóxido de Carbono/química , Concentración de Iones de Hidrógeno , Residuos Industriales , Metano/química , Methanomicrobiaceae/metabolismo , ARN Ribosómico 16S/genética , ARN Ribosómico 16S/metabolismo , Aguas del Alcantarillado/química , Factores de Tiempo , Treponema/metabolismo , Eliminación de Residuos LíquidosRESUMEN
This clinical study was conducted to quantify cultivable bacteria and endotoxin in root canals with post-treatment apical periodontitis by correlating their levels with clinical features and to evaluate the effect of chemo-mechanical preparation (CMP) with 2 % chlorhexidine gel + 17 % EDTA on bacterial and endotoxin removal/elimination. Moreover, target strict Gram-negative anaerobic bacteria were investigated by polymerase chain reaction (PCR). Fifteen teeth with post-treatment apical periodontitis were sampled before (s1) and after (s2) CMP. Culture techniques determined the number of colony-forming units (CFU). PCR (16S rDNA) and limulus amebocyte lysate (LAL) assay were used for bacterial and endotoxin detection, respectively. Prevotella nigrescens (4/15), Prevotella intermedia (2/15), and Tannerella forsythia (2/15) were the most frequently detected species. Endotoxin was recovered in 100 % of the samples. At s1, bacteria and endotoxin were detected at a median value of 5.14 × 10(3) CFU/mL and 3.96 EU/mL, respectively. Higher levels of endotoxin were related to a larger size of radiolucent area (>5 mm) (p < 0.05). CMP was more effective in reducing bacteria (99.61 %) than endotoxin (60.6 %) (both p < 0.05). Our findings indicated that the levels of endotoxin found in infected root canals were related to a larger size of radiolucent area in the periapical region. Moreover, CMP was effective in reducing both bacterial and endotoxin contents in post-treatment apical periodontitis.
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Carga Bacteriana/métodos , Clorhexidina/farmacología , Endotoxinas/metabolismo , Periodontitis Periapical/tratamiento farmacológico , Antibacterianos/farmacología , Técnicas de Tipificación Bacteriana/métodos , Cavidad Pulpar/metabolismo , Cavidad Pulpar/microbiología , Ácido Edético/farmacología , Humanos , Viabilidad Microbiana , Periodontitis Periapical/metabolismo , Periodontitis Periapical/microbiología , Reacción en Cadena de la Polimerasa/métodos , Prevotella/genética , Prevotella/crecimiento & desarrollo , Prevotella/aislamiento & purificación , Prevotella/metabolismo , ARN Bacteriano/análisis , ARN Bacteriano/genética , ARN Ribosómico 16S/análisis , ARN Ribosómico 16S/genética , Resultado del Tratamiento , Treponema/genética , Treponema/crecimiento & desarrollo , Treponema/aislamiento & purificación , Treponema/metabolismoRESUMEN
We have completed a bioinformatic analysis of the hydrogenases encoded in the genomes of three termite gut treponeme isolates: hydrogenotrophic, homoacetogenic Treponema primitia strains ZAS-1 and ZAS-2, and the hydrogen-producing, sugar-fermenting Treponema azotonutricium ZAS-9. H(2) is an important free intermediate in the breakdown of wood by termite gut microbial communities, reaching concentrations in some species exceeding those measured for any other biological system. The spirochetes encoded 4, 8, and 5 [FeFe] hydrogenase-like proteins, identified by their H domains, respectively, but no other recognizable hydrogenases. The [FeFe] hydrogenases represented many sequence families previously proposed in an analysis of termite gut metagenomic data. Each strain encoded both putative [FeFe] hydrogenase enzymes and evolutionarily related hydrogen sensor/transducer proteins likely involved in phosphorelay or methylation pathways, and possibly even chemotaxis. A new family of [FeFe] hydrogenases (FDH-Linked) is proposed that may form a multimeric complex with formate dehydrogenase to provide reducing equivalents for reductive acetogenesis in T. primitia. The many and diverse [FeFe] hydrogenase-like proteins encoded within the sequenced genomes of the termite gut treponemes has enabled the discovery of a putative new class of [FeFe] hydrogenase proteins potentially involved in acetogenesis and furthered present understanding of many families, including sensory, of H domain proteins beyond what was possible through the use of fragmentary termite gut metagenome sequence data alone, from which they were initially defined.
Asunto(s)
Proteínas Bacterianas/genética , Hidrogenasas/genética , Isópteros/microbiología , Treponema/genética , Animales , Formiato Deshidrogenasas/genética , Formiato Deshidrogenasas/metabolismo , Tracto Gastrointestinal/metabolismo , Tracto Gastrointestinal/microbiología , Hidrógeno/metabolismo , Isópteros/metabolismo , Datos de Secuencia Molecular , Familia de Multigenes , Filogenia , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Treponema/clasificación , Treponema/enzimología , Treponema/metabolismoRESUMEN
The hindguts of wood-feeding termites typically contain hundreds of microbial species. Together with their insect host, these gut microbes degrade lignocellulose into usable catabolites. Although past research revealed many facets of the stepwise flow of metabolites in this scheme, not much is known about the breadth of interactions occurring between termite-gut microbes. Most of these microbes are thought to depend on, and to have co-speciated with, their host and each other for millions of years. In this study, we explored the interactions of two spirochetes previously isolated from the very same termite species. As hydrogen (H(2)) is the central free intermediate in termite-gut lignocellulose digestion, we focused on interactions between two closely related termite-gut spirochetes possessing complementary H(2) physiologies: one produces H(2), while the other consumes it. In vitro, these two Treponema species markedly enhanced each other's growth. RNA sequencing resolved the transcriptomes of these two closely related organisms, revealing that co-cultivation causes comprehensive changes in global gene expression. The expression of well over a 100 genes in each species was changed >twofold, with over a dozen changed >10-fold. Several changes implicating synergistic cross-feeding of known metabolites were validated in vitro. Additionally, certain activities beneficial to the host were preferentially expressed during consortial growth. However, the majority of changes in gene expression are not yet understandable, but indicate a broad, comprehensive and mutualistic interaction between these closely related, co-resident gut symbionts. The results suggest that staggeringly intricate networks of metabolic and gene interactions drive lignocellulose degradation and co-evolution of termite gut microbiota.
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Tracto Gastrointestinal/microbiología , Hidrógeno/metabolismo , Isópteros/microbiología , Simbiosis , Treponema/metabolismo , Animales , Evolución Biológica , Técnicas de Cocultivo , Perfilación de la Expresión Génica , Genes Bacterianos , Lignina/metabolismo , Datos de Secuencia Molecular , ARN Bacteriano/genética , Análisis de Secuencia de ARN , Treponema/genética , Treponema/crecimiento & desarrolloRESUMEN
Treponema spp. are a commonly detected bacterial group in the rumen that are involved in the degradation of soluble fibers. In this study, a ruminal Treponema group-specific PCR primer targeting the 16S rRNA gene was designed and used to assess the phylogenetic diversity and diet association of this group in sheep rumen. Total DNA was extracted from rumen digesta of three sheep fed a diet based on alfalfa/orchardgrass hay or concentrate. The real-time PCR quantification indicated that the relative abundance of the Treponema group in the total rumen bacteria was as high as 1.05%, while the known species Treponema bryantii accounted for only 0.02%. Fingerprints of the Treponema community determined by 16S rDNA-targeted denaturing gradient gel electrophoresis (DGGE) analysis tended to differ among the diets. Principal component analysis of the DGGE profiles distinguished those Treponema associated with either the hay or the concentrate diets. Analysis of a Treponema 16S rRNA gene clone library showed phylogenetically distinct operational taxonomic units for a specific dietary condition, and significant (P=0.001) differences in community composition were observed among clone libraries constructed from each dietary regimen. The majority of clones (75.4%) had <97% sequence similarity with known Treponema. These results suggest the predominance of uncultured Treponema that appear to have distinct members related to the digestion of either hay or concentrate diet.
Asunto(s)
Biodiversidad , Rumen/microbiología , Treponema/clasificación , Treponema/metabolismo , Animales , Análisis por Conglomerados , Dermatoglifia del ADN , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Ribosómico/química , ADN Ribosómico/genética , Dieta/métodos , Electroforesis en Gel de Poliacrilamida , Datos de Secuencia Molecular , Desnaturalización de Ácido Nucleico , Filogenia , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Ovinos/microbiología , Treponema/genética , Treponema/aislamiento & purificaciónRESUMEN
During studies on fructan degradation in the rumen, a Treponema-like bacterium able to utilize Timothy grass fructan, commercial inulin and sucrose as the sole carbon source was recovered from sheep rumen. At least two different fructanolytic enzymes were identified in cell-free extracts of the isolated bacterium. Characterization of the strain by a polyphasic approach indicated that it can be regarded as a representative of a new bacterial species within the genus Treponema. Electron microscopy showed that the bacterium exhibited all of the features typical of spirochetes. The helical cells measured 5.4-11.5 microm x 0.42-0.51 microm and possessed up to seven regular coils. The bacterium utilized various plant mono- and disaccharides as fermentable substrates. Formate, acetate and ethanol in a molar ratio of 16 : 10 : 1 were the end products of glucose fermentation. The major cellular fatty acids were C(13:0), C(14:0), C(14:1), C(15:0), C(15:1) and C(16:0). The nearly complete 16S rRNA gene sequence was obtained, and phylogenetic analysis of the 16S rRNA gene showed the highest similarity to rumen Treponema strain CA. We propose the name Treponema zioleckii sp. nov. for this novel rumen spirochete with strain kT as the type strain.
Asunto(s)
Fructanos/metabolismo , Rumen/microbiología , Ovinos/microbiología , Treponema/aislamiento & purificación , Treponema/metabolismo , Animales , ADN Bacteriano/genética , ADN Ribosómico/genética , Fermentación , Datos de Secuencia Molecular , Filogenia , ARN Ribosómico 16S/genética , Treponema/clasificación , Treponema/citología , Treponema/genéticaRESUMEN
The class of Spirochetes comprises a wide array of clinically important pathogens, including Treponema pallidum causing syphilis as well as Borrelia burgdorferi, the agent of Lyme disease (LD). Diseases caused by spirochetes are characterized by specific sequelae of host reactions, and also by characteristic antibody response patterns. Over the last decades, research on the interaction of spirochetes with the host's immune system had a strong emphasis on outer membrane lipoproteins. In fact, these structures have been convincingly shown to activate immune cells via CD14 and Toll-like receptor (TLR)-2, and recent data also indicate an interaction with lipopolysaccharide (LPS)-binding protein (LBP). In particular, the interaction of B. burgdorferi with TLR-2 could not only be demonstrated in mice, but was also supported by data showing that genetic variants of TLR-2 in humans influenced the clinical course of LD. However, there is increasing evidence that next to lipoproteins, glycolipids may also play an important role in responses of the immune system towards spirochetes. Diacylglycerol-containing glycolipids exhibiting similarities with lipoteichoic acid (LTA) of Gram-positive bacteria have been demonstrated in various Treponema species, whereas LPS-like glycolipids have been shown to be present in Leptospira. Treponema glycolipids, comparably to lipoproteins and LTA, interact with LBP, CD14 and TLRs. In contrast, complex glycolipids of high molecular weight could not be demonstrated in Borrelia, whereas these bacteria exhibit a number of unique low molecular weight glycolipids. Some of these glycolipids cause strong immediate immune responses, while others appear to be potent antigens for induction of an adaptive immune response. This review summarizes data obtained so far on amphiphilic and hydrophobic molecules from spirochetes regarding structure and influence on innate as well as adaptive immune responses.
Asunto(s)
Glucolípidos/metabolismo , Sistema Inmunológico , Inmunidad Innata , Lipoproteínas/metabolismo , Enfermedad de Lyme/microbiología , Proteínas de Fase Aguda/metabolismo , Animales , Borrelia burgdorferi/metabolismo , Proteínas Portadoras/metabolismo , Diglicéridos/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Células Asesinas Naturales/inmunología , Leptospira/metabolismo , Glicoproteínas de Membrana/metabolismo , Modelos Biológicos , Treponema/metabolismoRESUMEN
Oral spirochetes include enormously heterogeneous Treponema species, and some have been implicated in the etiology of periodontitis. In this study, we characterized highly conserved surface proteins in four representative oral spirochetes (Treponema denticola, T. lecithinolyticum, T. maltophilum, and T. socranskii subsp. socranskii) that are homologs of T. pallidum Tp92, with opsonophagocytic potential and protective capacity against syphilis. Tp92 homologs of oral spirochetes had predicted signal peptides (20 to 31 amino acids) and molecular masses of 88 to 92 kDa for mature proteins. They showed amino acid sequence identities of 37.9 to 49.3% and similarities of 54.5 to 66.9% to Tp92. The sequence identities and similarities of Tp92 homologs of oral treponemes to one another were 41.6 to 71.6% and 59.9 to 85.6%, respectively. The tp92 gene homologs were successfully expressed in Escherichia coli, and the recombinant proteins were capable of binding to KB cells, an epithelial cell line, and inhibited the binding of the whole bacteria to the cells. Antiserum (the immunoglobulin G fraction) raised against a recombinant form of the T. denticola Tp92 homolog cross-reacted with homologs from three other species of treponemes. The Tp92 homologs stimulated various factors involved in inflammation and osteoclastogenesis, like interleukin-1beta (IL-1beta), tumor necrosis factor alpha, IL-6, prostaglandin E(2), and matrix metalloproteinase 9, in host cells like monocytes and fibroblasts. Our results demonstrate that Tp92 homologs of oral spirochetes are highly conserved and may play an important role in cell attachment, inflammation, and tissue destruction. The coexistence of various Treponema species in a single periodontal pocket and, therefore, the accumulation of multiple Tp92 homologs may amplify the pathological effect in periodontitis.
Asunto(s)
Adhesinas Bacterianas/metabolismo , Antígenos de Superficie/metabolismo , Proteínas Bacterianas/metabolismo , Inflamación/metabolismo , Osteoclastos/metabolismo , Treponema/metabolismo , Adhesinas Bacterianas/genética , Adhesinas Bacterianas/inmunología , Secuencia de Aminoácidos , Anticuerpos Antibacterianos/inmunología , Antígenos de Superficie/química , Antígenos de Superficie/genética , Antígenos de Superficie/inmunología , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/inmunología , Línea Celular , Clonación Molecular , Secuencia Conservada , Células Epiteliales/metabolismo , Regulación Bacteriana de la Expresión Génica , Humanos , Inmunoglobulina G/inmunología , Datos de Secuencia Molecular , Unión Proteica , Treponema/genética , Regulación hacia ArribaRESUMEN
Treponema lecithinolyticum is associated with periodontitis and endodontic infections. As a critical early step in the infection process, fibronectin-binding protein (Fbp) is known to be involved in the adhesion of bacteria to cell surfaces for colonization and, hence, is considered to be a virulence factor. In this study, we identified an Fbp from the T. lecithinolyticum cell surface with a molecular mass of about 52 kDa by using 2-dimensional gel electrophoresis followed by a ligand binding assay. As T. lecithinolyticum is capable of binding to soluble and immobilized fibronectin, this Fbp may contribute to bacterial attachment to host cells.
Asunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Electroforesis en Gel Bidimensional/métodos , Fibronectinas/metabolismo , Treponema/metabolismo , Secuencia de Aminoácidos , Humanos , Ligandos , Datos de Secuencia Molecular , Unión ProteicaRESUMEN
OMIZ-W68, a chemically defined medium that contains no long-chain fatty acids and yet supports in vitro proliferation of a wide range of fastidious oral anaerobes, is described. The type strains of Treponema denticola, Treponema lecithinolyticum, Treponema maltophilum, Treponema pectinovorum, Treponema socranskii, and an as yet unpublished canine Treponema species could be propagated indefinitely in this medium with sugar supplements for the saccharolytic species. Analysis of the cellular fatty acids (CFA) of these treponemes by gas chromatography demonstrated the synthesis of C14, C15, C16, and C17 fatty acids (linear-, iso-, and anteiso-forms) in various proportions, but neither hydroxy- nor unsaturated fatty acids. However, between 0% and 40% of the eluted material could not be identified. The proportions of CFAs differed not only between species but also between the eight strains of Treponema denticola investigated. Replacing OMIZ-W68 by a derivative minimal essential medium (OMIZ-M/TDCDK) developed for Treponema denticola had little effect on the CFA profiles. In contrast, the CFA profiles of treponemes grown in OMIZ-W68 showed at best minor similarity to the strains from the Moore library of the Virginia Polytechnic Institute, which had been grown in media containing serum, peptones, and yeast extract.