Coaggregation-Mediated Interactions of Streptococci and Actinomyces Detected in Initial Human Dental Plaque

ABSTRACT Streptococci and actinomyces that initiate colonization of the tooth surface frequently coaggregate with each other as well as with other oral bacteria. These observations have led to the hypothesis that interbacterial adhesion influences spatiotemporal development of plaque. To assess the role of such interactions in oral biofilm formation in vivo, antibodies directed against bacterial surface components that mediate coaggregation interactions were used as direct immunofluorescent probes in conjunction with laser confocal microscopy to determine the distribution and spatial arrangement of bacteria within intact human plaque formed on retrievable enamel chips. In intrageneric coaggregation, streptococci such as Streptococcus gordonii DL1 recognize receptor polysaccharides (RPS) borne on other streptococci such as Streptococcus oralis 34. To define potentially interactive subsets of streptococci in the developing plaque, an antibody against RPS (anti-RPS) was used together with an antibody against S. gordonii DL1 (anti-DL1). These antibodies reacted primarily with single cells in 4-h-old plaque and with mixed-species microcolonies in 8-h-old plaque. Anti-RPS-reactive bacteria frequently formed microcolonies with anti-DL1-reactive bacteria and with other bacteria distinguished by general nucleic acid stains. In intergeneric coaggregation between streptococci and actinomyces, type 2 fimbriae of actinomyces recognize RPS on the streptococci. Cells reactive with antibody against type 2 fimbriae of Actinomyces naeslundii T14V (anti-type-2) were much less frequent than either subset of streptococci. However, bacteria reactive with anti-type-2 were seen in intimate association with anti-RPS-reactive cells. These results are the first direct demonstration of coaggregation-mediated interactions during initial plaque accumulation in vivo. Further, these results demonstrate the spatiotemporal development and prevalence of mixed-species communities in early dental plaque.

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Probing of Microbial Biofilm Communities for Coadhesion Partners

Applied and Environmental Microbiology ◽

10.1128/aem.01826-14 ◽

2014 ◽

Vol 80 (21) ◽

pp. 6583-6590 ◽

Cited By ~ 10

Author(s):

Stefan Ruhl ◽

Andreas Eidt ◽

Holger Melzl ◽

Udo Reischl ◽

John O. Cisar

Keyword(s):

Dental Plaque ◽

Solid Phase ◽

Screening Method ◽

Bacterial Strains ◽

Mixed Species ◽

Simple Method ◽

Content Type ◽

Glycan Receptors ◽

Fimbrial Adhesin

ABSTRACTInvestigations of interbacterial adhesion in dental plaque development are currently limited by the lack of a convenient assay to screen the multitude of species present in oral biofilms. To overcome this limitation, we developed a solid-phase fluorescence-based screening method to detect and identify coadhesive partner organisms in mixed-species biofilms. The applicability of this method was demonstrated using coaggregating strains of type 2 fimbrial adhesin-bearing actinomyces and receptor polysaccharide (RPS)-bearing streptococci. Specific adhesin/receptor-mediated coadhesion was detected by overlaying bacterial strains immobilized to a nitrocellulose membrane with a suspended, fluorescein-labeled bacterial partner strain. Coadhesion was comparable regardless of which cell type was labeled and which was immobilized. Formaldehyde treatment of bacteria, either in suspension or immobilized on nitrocellulose, abolished actinomyces type 2 fimbrial adhesin but not streptococcal RPS function, thereby providing a simple method for assigning complementary adhesins and glycan receptors to members of a coadhering pair. The method's broader applicability was shown by overlaying colony lifts of dental plaque biofilm cultures with fluorescein-labeled strains of type 2 fimbriatedActinomyces naeslundiior RPS-bearingStreptococcus oralis. Prominent coadhesion partners included not only streptococci and actinomyces, as expected, but also other bacteria not identified in previous coaggregation studies, such as adhesin- or receptor-bearing strains ofNeisseria pharyngitis,Rothia dentocariosa, andKingella oralis. The ability to comprehensively screen complex microbial communities for coadhesion partners of specific microorganisms opens a new approach in studies of dental plaque and other mixed-species biofilms.

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Aggregative Behavior of Bacteria Isolated from Canine Dental Plaque

Applied and Environmental Microbiology ◽

10.1128/aem.01060-05 ◽

2006 ◽

Vol 72 (8) ◽

pp. 5211-5217 ◽

Cited By ~ 13

Author(s):

David R. Elliott ◽

Michael Wilson ◽

Catherine M. F. Buckley ◽

David A. Spratt

Keyword(s):

Electron Microscopy ◽

Transmission Electron Microscopy ◽

Dental Plaque ◽

Oral Bacteria ◽

Surface Structures ◽

Aggregative Behavior ◽

Transmission Electron

ABSTRACT Interbacterial adhesion of bacteria isolated from canine dental plaque was assessed by performing a visual coaggregation assay. Using conditions mimicking those likely to be encountered in vivo, the entire cultivable plaque microbiota from a single dog was assessed, and eight (6.7%) unique coaggregation interactions were detected for 120 crosses. Transmission electron microscopy was used to visualize several of the bacteria in isolation and as coaggregates, which revealed surface structures that may be involved in adhesion and coaggregation. The results of this study indicate that the prevalence of coaggregating pairs of dental plaque bacteria in dogs is similar to the prevalence of coaggregating pairs of dental plaque bacteria reported in humans. In addition, genera found in both hosts generally exhibited similar coaggregation reactions; however, autoaggregation was found to be more common among oral bacteria isolated from dogs.

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Single-Cell Genomics and the Oral Microbiome

Journal of Dental Research ◽

10.1177/0022034520907380 ◽

2020 ◽

Vol 99 (6) ◽

pp. 613-620 ◽

Cited By ~ 2

Author(s):

M. Balachandran ◽

K.L. Cross ◽

M. Podar

Keyword(s):

Microbial Community ◽

Single Cell ◽

Single Cells ◽

Oral Bacteria ◽

Oral Microbiome ◽

Oral Microbiota ◽

Oral Diseases ◽

Interspecies Interactions ◽

Single Cell Genomics

The human oral cavity is one of the first environments where microbes have been discovered and studied since the dawn of microbiology. Nevertheless, approximately 200 types of bacteria from the oral microbiota have remained uncultured in the laboratory. Some are associated with a healthy oral microbial community, while others are linked to oral diseases, from dental caries to gum disease. Single-cell genomics has enabled inferences on the physiology, virulence, and evolution of such uncultured microorganisms and has further enabled isolation and cultivation of several novel oral bacteria, including the discovery of novel interspecies interactions. This review summarizes some of the more recent advances in this field, which is rapidly moving toward physiologic characterization of single cells and ultimately cultivation of the yet uncultured. A combination of traditional microbiological approaches with genomic-based physiologic predictions and isolation strategies may lead to the oral microbiome being the first complex microbial community to have all its members cultivable in the laboratory. Studying the biology of the individual microbes when in association with other members of the community, in controlled laboratory conditions and in vivo, should lead to a better understanding of oral dysbiosis and its prevention and reversion.

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Polymerase Chain Reaction for detection of Streptococcus mutans and Streptococcus sobrinus in dental plaque of children from Cartagena, Colombia

Colombia Medica ◽

10.25100/cm.v42i4.943 ◽

2011 ◽

pp. 430-437 ◽

Cited By ~ 1

Author(s):

Luis Eduardo Carmona ◽

Niradiz Reyes ◽

Farith González

Keyword(s):

Streptococcus Mutans ◽

Dental Plaque ◽

Statistical Significance ◽

Study Groups ◽

Oral Bacteria ◽

Tooth Surface ◽

Streptococcus Sobrinus ◽

Exact Test ◽

Carious Lesions ◽

The Difference

Objectives: To detect the presence of Streptococcus mutans and Streptococcus sobrinus in dental plaque of children from Cartagena and correlate it to dental caries precavity stages, applying a standardized PCR-based technique for epidemiological purposes. Methods: Descriptive study using a non-probabilistic sample of 50 children between 3 and 5 years of age, preschoolers from a Caribbean population in Colombia. Criteria for selection were that children should exhibit plaque accumulations on the surface of the cervical margins of the rearmost molars, and placed in one of two study groups: carious lesions and sound surfaces. Dental plaque samples from both groups were subjected to molecular analysis and statistical analysis was applied to determine the difference between the two groups using the frequencies of presence of S. mutans, S. sobrinus or both in the two groups applying Fisher’s exact test for association between the presence of microorganisms and the state of the tooth surface from where the dental plaque was taken. Results: The frequency of S. mutans in carious lesions was 76% and 24% in healthy surfaces. The frequency of S. sobrinus in carious lesions was 81.9% and 18.1% in caries-free surfaces. There was statistical significance between the presence of S. mutans and the presence of caries (p=0.001) and between the presence of S. sobrinus (p=0.02) and the presence of caries. There was no statistical significance between the presence of caries and the simultaneous presence of both microorganisms (p=0.08). Conclusions: The presence of S. mutans and S. sobrinus in dental plaque samples is highly prevalent and associated to non cavitated carious lesions, being the molecular identification of these microorganisms by PCR a sensitive, fast, and easy to use detection method for the mutans group of oral bacteria.

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Short chain fatty acids induced the type 1 and type 2 fimbrillin-dependent and fimbrillin-independent initial attachment and colonization of Actinomyces oris monoculture but not coculture with streptococci

BMC Microbiology ◽

10.1186/s12866-020-01976-4 ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Itaru Suzuki ◽

Takehiko Shimizu ◽

Hidenobu Senpuku

Keyword(s):

Fatty Acids ◽

Short Chain Fatty Acids ◽

Human Saliva ◽

Oral Bacteria ◽

Short Chain ◽

Tooth Surface ◽

Initial Attachment ◽

Chain Fatty Acids

Abstract Background Actinomyces oris is an early colonizer and has two types of fimbriae on its cell surface, type 1 fimbriae (FimP and FimQ) and type 2 fimbriae (FimA and FimB), which contribute to the attachment and coaggregation with other bacteria and the formation of biofilm on the tooth surface, respectively. Short-chain fatty acids (SCFAs) are metabolic products of oral bacteria including A. oris and regulate pH in dental plaques. To clarify the relationship between SCFAs and fimbrillins, effects of SCFAs on the initial attachment and colonization (INAC) assay using A. oris wild type and fimbriae mutants was investigated. INAC assays using A. oris MG1 strain cells were performed with SCFAs (acetic, butyric, propionic, valeric and lactic acids) or a mixture of them on human saliva-coated 6-well plates incubated in TSB with 0.25% sucrose for 1 h. The INAC was assessed by staining live and dead cells that were visualized with a confocal microscope. Results Among the SCFAs, acetic, butyric and propionic acids and a mixture of acetic, butyric and propionic acids induced the type 1 and type 2 fimbriae-dependent and independent INAC by live A. oris, but these cells did not interact with streptococci. The main effects might be dependent on the levels of the non-ionized acid forms of the SCFAs in acidic stress conditions. GroEL was also found to be a contributor to the FimA-independent INAC by live A. oris cells stimulated with non-ionized acid. Conclusion SCFAs affect the INAC-associated activities of the A. oris fimbrillins and non-fimbrillins during ionized and non-ionized acid formations in the form of co-culturing with other bacteria in the dental plaque but not impact the interaction of A. oris with streptococci.

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New Fine Structure within the Adult Hematopoietic Stem Cell Compartment Revealed by Longterm Analysis of Mice Repopulated with Single Cells or Their In Vitro Generated Clonal Progeny.

Blood ◽

10.1182/blood.v106.11.1720.1720 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1720-1720

Author(s):

Brad Dykstra ◽

David Kent ◽

Lindsay McCaffrey ◽

Kristin Lyons ◽

Merete Kristiansen ◽

...

Keyword(s):

Stem Cell ◽

Hematopoietic Stem Cell ◽

Single Cells ◽

Myeloid Cells ◽

Hematopoietic Stem ◽

Type 3

Abstract Assessments of hematopoietic stem cell (HSC) repopulating activity in vivo have historically relied on calculated average longterm (12–16 wk) progeny outputs using non-purified transplants, thereby precluding definitive clonal assignments of donor-derived cells. Viral marking circumvents this problem, but has not been used for large scale surveys. Heterogeneity observed in the repopulation patterns has generally been inferred to reflect stochastic processes. We now report the in vivo repopulation kinetics of 89 individual longterm repopulating cells (LTRCs) before (n=49) and after (n=40) 4 days of clonal growth in vitro. LTRCs were defined here as cells whose WBC progeny could be detected at levels of ≥1% for at least 16 wk in sublethally irradiated Ly5-congenic W41/W41 hosts. Recipients were transplanted with either freshly isolated, single lin−Rho−SP LTRCs or 4-day clones generated from similar cells in serum-free cultures (+ 300 ng/ml SF, 20 ng/ml IL-11 & 1ng/ml Flt3-L). 4, 8, 12, 16, and 24 wk post-transplant, blood samples were stained for donor-derived B, T, and myeloid cells using a procedure that identifies donor/recipient doublets and Ly6g/Mac1low cells (which have features of lymphoid rather than myeloid WBCs) to exclude false-positive myeloid events. Four distinct patterns of repopulation were revealed. Type 1 showed a delayed production of predominantly myeloid WBCs (low or undetectable before 12 wk) that increased progressively (reaching 0.4–15% of all WBCs by 16 wk). Type 2 showed a robust multilineage repopulation that remained stable or increased over time (6–84% of all WBCs at 16 wk). Type 3 also showed an initially robust pattern of multilineage repopulation (peak numbers of WBCs at 8–12 wk and 1–51% at 16 wk), but the contribution of donor-derived myeloid cells was transient (<0.5% by 16 wk). Type 4 showed a lymphoid-restricted pattern (myeloid contribution <0.5% at all time points), with repopulation levels peaking at 8 wk and decreasing thereafter (1–22% at 16 wk). Persisting granulopoiesis, indicated by a high proportion of donor-derived cells in the Ly6g/Mac1+SSChi population at 16–24 wk, clearly distinguished the type 1 and 2 patterns from types 3 and 4 which showed near or complete cessation of donor-derived granulopoiesis beyond 12 wk. Preliminary secondary transplant experiments show that donor-derived LTRCs (with and without longterm granulopoietic activity) were exclusively generated in primary recipients with type 1 and 2 repopulation patterns. Amongst the freshly isolated LTRCs, 18% (9/49) were type 1, 41% (20/49) were type 2, 22% (11/49) were type 3, and 18% (9/49) were type 4. In contrast, 4-day clones derived from cells of the same phenotype and containing LTRC activity showed a marked decrease in type 1 and type 2 activity with a corresponding increase in type 3 and type 4 activity: type 1 = 5% (2/41), type 2 = 18% (7/40), type 3 = 28% (11/40) and type 4 = 50% (20/40). Collectively, these data identify a new hierarchy of four biologically discrete states within the compartment of cells currently defined as LTRCs. Proliferation of LTRCs either in vitro or in vivo appears to induce an irreversible transition from one state to another (from Type 1 to 2 to 3 to 4), suggesting the existence of intrinsic molecular correlates for each of these states and specific mechanisms that underlie their sequential appearance.

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A polymicrobial biofilm model for testing the antimicrobial potential of a nisin-biogel for canine periodontal disease control

BMC Veterinary Research ◽

10.1186/s12917-020-02646-3 ◽

2020 ◽

Vol 16 (1) ◽

Author(s):

Eva Cunha ◽

Sandra Rebelo ◽

Carla Carneiro ◽

Luís Tavares ◽

Luís Miguel Carreira ◽

...

Keyword(s):

Periodontal Disease ◽

Guar Gum ◽

Bacterial Species ◽

Density Measurement ◽

Oral Bacteria ◽

Tooth Surface ◽

Pd Control ◽

Biofilm Model

Abstract Background Periodontal disease (PD) in dogs is prompted by the establishment of a polymicrobial biofilm at the tooth surface and a subsequent host inflammatory response. Several strategies may be used for PD control, including dental hygiene home care procedures, like toothbrushing, special diet and chew toys that reduce dental plaque accumulation, or professional periodontal treatments. Aiming at PD control, a biogel composed by nisin and guar-gum was previously developed. This work aimed to establish an in vitro model mimicking the PD-associated biofilms and to evaluate the nisin-biogel inhibitory activity against this polymicrobial biofilm by determining its Minimum Biofilm Inhibitory (MBIC) and Eradication Concentrations (MBEC). Bacterial species tested included Neisseria zoodegmatis CCUG 52598T, Corynebacterium canis CCUG 58627T, Porphyromonas cangingivalis DSMZ VPB 4874, Peptostreptococcus canis CCUG 57081 and an Enterococcus faecalis isolate belonging to a collection of oral bacteria obtained from dogs with PD. Before establishing the biofilm, coaggregation between species was determined by optical density measurement after 2 and 24 hours. Nisin-biogel MBIC and MBEC values regarding the polymicrobial biofilm were determined using a modified version of the Calgary biofilm pin lid device, after confirming the presence of the five bacterial species by Fluorescent In Situ Hybridization. Results Only 40% of the bacterial dual suspensions were able to coaggregate at 2 hours, but all species tested exhibited a coaggregation percentage higher than 30% at 24 hours. It was possible to establish a 48 h polymicrobial biofilm model composed by the five bacterial species selected. This model was used to determine nisin-biogel MBIC (26.39 ± 5.89 µg/mL) and MBEC (62.5 ± 27.73 µg/mL) values. Conclusions Our results showed that the nisin-biogel can inhibit and eradicate PD multispecies biofilms. As this in vitro model mimics an in vivo periodontal polymicrobial biofilm, our results reinforce the potential of the application of nisin-biogel for canine PD control.

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