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PloS One 2016Live attenuated influenza vaccine (LAIV) has demonstrated varying levels of efficacy against seasonal influenza; however, LAIV may be used as a tool to measure...
Live attenuated influenza vaccine (LAIV) has demonstrated varying levels of efficacy against seasonal influenza; however, LAIV may be used as a tool to measure interactions between the human microbiome and a live, replicating virus. To increase our knowledge of this interaction, we measured changes to the nasal microbiome in subjects who received LAIV to determine if associations between influenza-specific IgA production and the nasal microbiome exist after immunization with a live virus vaccine. The anterior nares of 47 healthy subjects were swabbed pre- (Day 0) and post- (Days 7 and 28) LAIV administration, and nasal washes were conducted on Days 0 and 28. We performed next-generation sequencing on amplified 16s rRNA genes and measured mucosal influenza-specific IgA titers via enzyme-linked immunosorbent assay (ELISA). A significant increase in alpha diversity was identified (Observed, CHAO, and ACE) between Days 7 vs 0 (p-values = 0.017, 0.005, 0.005, respectively) and between Days 28 vs 0 (p-values = 0.054, 0.030, 0.050, respectively). Several significant associations between the presence of different microbial species, including Lactobacillus helveticus, Prevotella melaninogenica, Streptococcus infantis, Veillonella dispar, and Bacteroides ovatus, and influenza-specific H1 and H3 IgA antibody response were demonstrated. These data suggest that LAIV alters the nasal microbiome, allowing several less-abundant OTUs to establish a community niche. Additionally, specific alterations in the nasal microbiome are significantly associated with variations in influenza-specific IgA antibody production and could be clinically relevant.
Topics: Adolescent; Adult; Antibodies, Viral; Bacteria; Female; Hemagglutinin Glycoproteins, Influenza Virus; Humans; Immunoglobulin A; Influenza A virus; Influenza Vaccines; Influenza, Human; Male; Microbiota; Nasal Cavity; Vaccines, Attenuated; Young Adult
PubMed: 27643883
DOI: 10.1371/journal.pone.0162803 -
Clinical Oral Implants Research Jul 2016Staphylococcus spp. are postulated to play a role in peri-implantitis. This study aimed to develop a "submucosal" in vitro biofilm model, by integrating two...
OBJECTIVES
Staphylococcus spp. are postulated to play a role in peri-implantitis. This study aimed to develop a "submucosal" in vitro biofilm model, by integrating two staphylococci into its composition.
MATERIALS AND METHODS
The standard "subgingival" biofilm contained Actinomyces oris, Fusobacterium nucleatum, Streptococcus oralis, Veillonella dispar, Campylobacter rectus, Prevotella intermedia, Streptococcus anginosus, Porphyromonas gingivalis, Tannerella forsythia and Treponema denticola, and was further supplemented with Staphyoccous aureus and/or Staphylococcus epidermidis. Biofilms were grown anaerobically on hydroxyapatite or titanium discs and harvested after 64 h for real-time polymerase chain reaction, to determine their composition. Confocal laser scanning microscopy and fluorescence in situ hybridization were used for identifying the two staphylococci within the biofilm.
RESULTS
Both staphylococci established within the biofilms when added separately. However, when added together, only S. aureus grew in high numbers, whereas S. epidermidis was reduced almost to the detection limit. Compared to the standard subgingival biofilm, addition of the two staphylococci had no impact on the qualitative or quantitative composition of the biofilm. When grown individually in the biofilm, S. epidermidis and S. aureus formed small distinctive clusters and it was confirmed that S. epidermidis was not able to grow in presence of S. aureus.
CONCLUSIONS
Staphyoccous aureus and S. epidermidis can be individually integrated into an oral biofilm grown on titanium, hence establishing a "submucosal" biofilm model for peri-implantitis. This model also revealed that S. aureus outcompetes S. epidermidis when grown together in the biofilm, which may explain the more frequent association of the former with peri-implantitis.
Topics: Biofilms; Gingiva; Humans; In Vitro Techniques; Models, Biological; Peri-Implantitis; Staphylococcus aureus; Staphylococcus epidermidis; Titanium
PubMed: 26461083
DOI: 10.1111/clr.12715 -
Applied and Environmental Microbiology Dec 2015Around a third of oral bacteria cannot be grown using conventional bacteriological culture media. Community profiling targeting 16S rRNA and shotgun metagenomics methods...
Around a third of oral bacteria cannot be grown using conventional bacteriological culture media. Community profiling targeting 16S rRNA and shotgun metagenomics methods have proved valuable in revealing the complexity of the oral bacterial community. Studies investigating the role of oral bacteria in health and disease require phenotypic characterizations that are possible only with live cultures. The aim of this study was to develop novel culture media and use an in vitro biofilm model to culture previously uncultured oral bacteria. Subgingival plaque samples collected from subjects with periodontitis were cultured on complex mucin-containing agar plates supplemented with proteose peptone (PPA), beef extract (BEA), or Gelysate (GA) as well as on fastidious anaerobe agar plus 5% horse blood (FAA). In vitro biofilms inoculated with the subgingival plaque samples and proteose peptone broth (PPB) as the growth medium were established using the Calgary biofilm device. Specific PCR primers were designed and validated for the previously uncultivated oral taxa Bacteroidetes bacteria HOT 365 and HOT 281, Lachnospiraceae bacteria HOT 100 and HOT 500, and Clostridiales bacterium HOT 093. All agar media were able to support the growth of 10 reference strains of oral bacteria. One previously uncultivated phylotype, Actinomyces sp. HOT 525, was cultivated on FAA. Of 93 previously uncultivated phylotypes found in the inocula, 26 were detected in in vitro-cultivated biofilms. Lachnospiraceae bacterium HOT 500 was successfully cultured from biofilm material harvested from PPA plates in coculture with Parvimonas micra or Veillonella dispar/parvula after colony hybridization-directed enrichment. The establishment of in vitro biofilms from oral inocula enables the cultivation of previously uncultured oral bacteria and provides source material for isolation in coculture.
Topics: Actinomyces; Bacteriological Techniques; Bacteroidetes; Biofilms; Clostridiales; Culture Media; Dental Plaque; Humans; In Situ Hybridization, Fluorescence; Microbiota; Molecular Sequence Data; Mouth; Periodontitis; RNA, Ribosomal, 16S
PubMed: 26407883
DOI: 10.1128/AEM.02156-15 -
Journal of Oral Microbiology 2015Bacterial invasion into pulps of primary teeth can lead to infection and premature tooth loss in children. This pilot study aimed to explore whether the microbiota of...
BACKGROUND AND OBJECTIVE
Bacterial invasion into pulps of primary teeth can lead to infection and premature tooth loss in children. This pilot study aimed to explore whether the microbiota of carious exposures of dental pulps resembles that of carious dentin or that of infected root canals.
DESIGN
Children with severe early childhood caries were studied. Children were consented and extent of caries, plaque, and gingivitis measured. Bacteria were sampled from carious lesion biofilms and vital carious exposures of pulps, and processed by anaerobic culture. Isolates were characterized from partial sequences of the 16S rRNA gene and identified by comparison with taxa in the Human Oral Microbiome Database (http://www.HOMD.org). The microbiotas of carious lesions and dental pulps were compared using univariate and multivariate approaches.
RESULTS
The microbiota of cariously exposed pulps was similar in composition to that of carious lesion biofilms except that fewer species/taxa were identified from pulps. The major taxa identified belonged to the phyla Firmicutes (mainly streptococci) and Actinobacteria (mainly Actinomyces species). Actinomyces and Selenomonas species were associated with carious lesions whereas Veillonella species, particularly Veillonella dispar was associated with pulps. Other bacteria detected in pulps included Streptococcus mutans, Parascardovia denticolens, Bifidobacterium longum, and several Lactobacillus and Actinomyces species. By principal, component analysis pulp microbiotas grouped together, whereas those in caries biofilms were widely dispersed.
CONCLUSIONS
We conclude that the microbiota of cariously exposed vital primary pulps is composed of a subset of species associated with carious lesions. Vital primary pulps had a dominant Firmicutes and Actinobacteria microbiota which contrasts with reports of endodontic infections which can harbor a gram-negative microbiota. The microbiota of exposed primary pulps may provide insight into bacterial species at the forefront of caries invasion in dentinal lesions that can invade into the pulp and the nature of species that need suppressing for successful pulp therapy.
PubMed: 25651832
DOI: 10.3402/jom.v7.25951 -
Virulence 2015Periodontal infection involves a complex interplay between oral biofilms, gingival tissues and cells of the immune system in a dynamic microenvironment. A humanized in...
Periodontal infection involves a complex interplay between oral biofilms, gingival tissues and cells of the immune system in a dynamic microenvironment. A humanized in vitro model that reduces the need for experimental animal models, while recapitulating key biological events in a periodontal pocket, would constitute a technical advancement in the study of periodontal disease. The aim of this study was to use a dynamic perfusion bioreactor in order to develop a gingival epithelial-fibroblast-monocyte organotypic co-culture on collagen sponges. An 11 species subgingival biofilm was used to challenge the generated tissue in the bioreactor for a period of 24 h. The histological and scanning electron microscopy analysis displayed an epithelial-like layer on the surface of the collagen sponge, supported by the underlying ingrowth of gingival fibroblasts, while monocytic cells were also found within the sponge mass. Bacterial quantification of the biofilm showed that in the presence of the organotypic tissue, the growth of selected biofilm species, especially Campylobacter rectus, Actinomyces oris, Streptococcus anginosus, Veillonella dispar, and Porphyromonas gingivalis, was suppressed, indicating a potential antimicrobial effect by the tissue. Multiplex immunoassay analysis of cytokine secretion showed that interleukin (IL)-1 β, IL-2, IL-4, and tumor necrosis factor (TNF)-α levels in cell culture supernatants were significantly up-regulated in presence of the biofilm, indicating a positive inflammatory response of the organotypic tissue to the biofilm challenge. In conclusion, this novel host-biofilm interaction organotypic model might resemble the periodontal pocket and have an important impact on the study of periodontal infections, by minimizing the need for the use of experimental animal models.
Topics: Biofilms; Bioreactors; Cell Line; Coculture Techniques; Collagen; Fibrocartilage; Gingiva; Humans; Interleukins; Organ Culture Techniques; Perfusion; Periodontal Pocket; Porphyromonas gingivalis; Tumor Necrosis Factor-alpha
PubMed: 25587671
DOI: 10.4161/21505594.2014.978721 -
Applied and Environmental Microbiology Jul 2014Indigenous oral bacteria in the tongue coating such as Veillonella have been identified as the main producers of hydrogen sulfide (H2S), one of the major components of...
Indigenous oral bacteria in the tongue coating such as Veillonella have been identified as the main producers of hydrogen sulfide (H2S), one of the major components of oral malodor. However, there is little information on the physiological properties of H2S production by oral Veillonella such as metabolic activity and oral environmental factors which may affect H2S production. Thus, in the present study, the H2S-producing activity of growing cells, resting cells, and cell extracts of oral Veillonella species and the effects of oral environmental factors, including pH and lactate, were investigated. Type strains of Veillonella atypica, Veillonella dispar, and Veillonella parvula were used. These Veillonella species produced H2S during growth in the presence of l-cysteine. Resting cells of these bacteria produced H2S from l-cysteine, and the cell extracts showed enzymatic activity to convert l-cysteine to H2S. H2S production by resting cells was higher at pH 6 to 7 and lower at pH 5. The presence of lactate markedly increased H2S production by resting cells (4.5- to 23.7-fold), while lactate had no effect on enzymatic activity in cell extracts. In addition to H2S, ammonia was produced in cell extracts of all the strains, indicating that H2S was produced by the catalysis of cystathionine γ-lyase (EC 4.4.1.1). Serine was also produced in cell extracts of V. atypica and V. parvula, suggesting the involvement of cystathionine β-synthase lyase (EC 4.2.1.22) in these strains. This study indicates that Veillonella produce H2S from l-cysteine and that their H2S production can be regulated by oral environmental factors, namely, pH and lactate.
Topics: Alanine; Ammonia; Cystathionine beta-Synthase; Cystathionine gamma-Lyase; Cysteine; Hydrogen Sulfide; Hydrogen-Ion Concentration; Lactic Acid; Serine; Sulfides; Veillonella
PubMed: 24795374
DOI: 10.1128/AEM.00606-14 -
Journal of Agricultural and Food... May 2014The antimicrobial effects of red wine and its inherent components on oral microbiota were studied by using a 5-species biofilm model of the supragingival plaque that...
The antimicrobial effects of red wine and its inherent components on oral microbiota were studied by using a 5-species biofilm model of the supragingival plaque that includes Actinomyces oris, Fusobacterium nucleatum, Streptococcus oralis, Streptococcus mutans and Veillonella dispar. Microbiological analysis (CFU counting and confocal laser scanning microscopy) of the biofilms after the application of red wine, dealcoholized red wine, and red wine extract solutions spiked or not with grape seed and inactive dry yeast extracts showed that the solutions spiked with seed extract were effective against F. nucleatum, S. oralis and A. oris. Also, red wine and dealcoholized wine had an antimicrobial effect against F. nucleatum and S. oralis. Additional experiments showed almost complete and early degradation of flavan-3-ol precursors [(+)-catechin and procyanidin B2] when incubating biofilms with the red wine extract. To our knowledge, this is the first study of antimicrobial properties of wine in an oral biofilm model.
Topics: Actinomyces; Anti-Bacterial Agents; Biofilms; Color; Dental Plaque; Fusobacterium nucleatum; Humans; Plant Extracts; Streptococcus; Veillonella; Vitis; Wine
PubMed: 24773294
DOI: 10.1021/jf501768p -
PloS One 2013The aim of this study was to investigate the impact of early colonizing species on the structure and the composition of the bacterial community developing in a...
The aim of this study was to investigate the impact of early colonizing species on the structure and the composition of the bacterial community developing in a subgingival 10-species biofilm model system. The model included Streptococcus oralis, Streptococcus anginosus, Actinomycesoris, Fusobacterium nucleatum subsp. nucleatum, Veillonella dispar, Campylobacter rectus, Prevotella intermedia, Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola. Based on literature, we considered Streptococcus oralis, Streptococcus anginosus, and Actinomyces oris as early colonizers and examined their role in the biofilms by either a delayed addition to the consortium, or by not inoculating at all the biofilms with these species. We quantitatively evaluated the resulting biofilms by real-time quantitative PCR and further compared the structures using confocal laser scanning microscopy following fluorescence in situ hybridisation. The absence of the early colonizers did not hinder biofilm formation. The biofilms reached the same total counts and developed to normal thickness. However, quantitative shifts in the abundances of individual species were observed. In the absence of streptococci, the overall biofilm structure appeared looser and more dispersed. Moreover, besides a significant increase of P. intermedia and a decrease of P. gingivalis , P. intermedia appeared to form filamented long chains that resembled streptococci. A. oris, although growing to significantly higher abundance in absence of streptococci, did not have a visible impact on the biofilms. Hence, in the absence of the early colonizers, there is a pronounced effect on P. intermedia and P. gingivalis that may cause distinct shifts in the structure of the biofilm. Streptococci possibly facilitate the establishment of P. gingivalis into subgingival biofilms, while in their absence P. intermedia became more dominant and forms elongated chains.
Topics: Actinomyces; Biofilms; Campylobacter rectus; Colony Count, Microbial; Fusobacterium nucleatum; Gingiva; Humans; In Situ Hybridization, Fluorescence; Microbial Consortia; Microscopy, Confocal; Porphyromonas gingivalis; Prevotella intermedia; Real-Time Polymerase Chain Reaction; Streptococcus; Streptococcus anginosus; Streptococcus oralis; Treponema denticola; Veillonella
PubMed: 24340084
DOI: 10.1371/journal.pone.0083090 -
FEMS Immunology and Medical Microbiology Aug 2010We demonstrate that live donor Veillonella dispar cells can transfer the conjugative transposon Tn916 to four different Streptococcus spp. recipients in a multispecies...
We demonstrate that live donor Veillonella dispar cells can transfer the conjugative transposon Tn916 to four different Streptococcus spp. recipients in a multispecies oral consortium growing as a biofilm in a constant depth film fermentor. Additionally, we demonstrate that purified V. dispar DNA can transform Streptococcus mitis to tetracycline resistance in this experimental system. These data show that transfer of conjugative transposon-encoded antibiotic resistance can occur by transformation in addition to conjugation in biofilms.
Topics: Anti-Bacterial Agents; Biofilms; Conjugation, Genetic; DNA Transposable Elements; DNA, Bacterial; Drug Resistance, Bacterial; Humans; Streptococcus mitis; Transformation, Bacterial; Veillonella
PubMed: 20337719
DOI: 10.1111/j.1574-695X.2010.00661.x -
Antimicrobial Agents and Chemotherapy Aug 2006Twelve out of 96 Veillonella spp. isolated from oral samples harbored tetracycline resistance genes. The most common resistance gene was tet(M). A tet(M)-positive...
Twelve out of 96 Veillonella spp. isolated from oral samples harbored tetracycline resistance genes. The most common resistance gene was tet(M). A tet(M)-positive Veillonella dispar strain was shown to transfer a Tn916-like element to four Streptococcus spp. by conjugation at a frequency of 5.2 x 10(-6) to 4.5 x 10(-5) per recipient.
Topics: Bacterial Proteins; Conjugation, Genetic; DNA Transposable Elements; DNA, Bacterial; Drug Resistance, Bacterial; Gene Transfer, Horizontal; Genes, Bacterial; Humans; Microbial Sensitivity Tests; Mouth; Streptococcus; Tetracycline Resistance; Veillonella
PubMed: 16870789
DOI: 10.1128/AAC.00217-06