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Development of novel tools for prevention and diagnosis of Porphyromonas gingivalis infection and periodontitis
Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
2015 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Periodontitis is a chronic infectious disease causing inflammation and destruction of tooth-supporting structures eventually leading to tooth loss. Periodontitis is induced by different subgingival microorganisms in combination with an excessive and dysregulated immune-inflammatory response against this microbial challenge. The pathogenicity of the periodontal biofilm is highly dependent upon some "keystone" species of which Porphyromonas gingivalis is considered to be one of the most important pathogens. P. gingivalis expresses a broad range of virulence factors including lipopolysaccharides (LPS), fimbriae, adhesins, hemagglutinins and an array of proteolytic enzymes. Among these factors, cysteine proteases (gingipains) are of special importance both for the bacterial survival/proliferation and for the pathological outcome. Gingipains consist of arginine-specific proteases (RgpA and RgpB) and lysine-specific protease (Kgp). The major aim of this thesis was to develop and test novel methods for diagnosis and prevention of P. gingivalis infection and periodontitis. In study I, host lymphocytes were stimulated with P. gingivalis to develop antibodies in vitro and the anti-P. gingivalis antibodies were used for immunodetection in clinical samples. The in vitro method of antibody production developed during this study could be used for an efficient real-time detection of P. gingivalis infection and periodontitis, while the attenuating effects of the antibodies suggest a role in passive immunization to prevent periodontitis and its associated diseases. In study II, we have elucidated the properties and antimicrobial effects of different lactobacillus species and the two-peptide bacteriocin NC8 αβ on P. gingivalis. NC8 αβ was found to be efficient against P. gingivalis through bacterial binding followed by permeabilization of the membranes. Liposomal systems were acquired to verify membrane permeabilization by NC8 αβ. The antimicrobial activity of NC8 αβ was found to be rapid, potent and instant. In conclusion, soluble or immobilized NC8 αβ bacteriocins may be used to prevent P. gingivalis colonization and subsequent pathogenicity, and thus supplement the host immune system against invading pathogens associated with periodontitis.

Place, publisher, year, edition, pages
Örebro: Örebro University , 2015. , 30 p.
National Category
Microbiology in the medical area
Research subject
Biomedicine
Identifiers
URN: urn:nbn:se:oru:diva-46712OAI: oai:DiVA.org:oru-46712DiVA: diva2:873301
Available from: 2015-11-24 Created: 2015-11-23 Last updated: 2015-12-21Bibliographically approved
List of papers
1. Antibodies produced in vitro in the detection of periodontal bacteria by using surface plasmon resonance analysis
Open this publication in new window or tab >>Antibodies produced in vitro in the detection of periodontal bacteria by using surface plasmon resonance analysis
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2015 (English)In: Clinical and Experimental Dental Research, ISSN 2057-4347, Vol. 1, no 1, 32-44 p.Article in journal (Refereed) Published
Abstract [en]

Porphyromonas gingivalis (P. gingivalis) is a major etiological agent associated with periodontitis. This study aims to develop antibodies to P. gingivalis in vitro for real-time detection of bacteria in clinical samples. Lymphocytes were isolated from whole blood of patient treated for periodontitis and were stimulated with P. gingivalis ATCC 33277. B-cell maturation to long-living antibody secreting-plasma cells was studied using flow cytometry and immunofluorescence staining. The antibodies developed in vitro were immobilized onto a CM-5 sensor chip of a biosensor to detect the presence of P. gingivalis in the gingival crevicular fluid of patients with periodontitis compared to periodontally healthy controls (n = 30). Surface plasmon resonance (SPR) analysis was performed to evaluate specific interactions of bacteria in samples with the immobilized antibodies. The results of SPR analysis were compared to the detection of P. gingivalis in the samples using DNA–DNA checkerboard hybridization technique. A clear and distinct change in lymphocyte morphology upon stimulation with P. gingivalis was observed. Anti-P. gingivalis antibodies secreted by CD38+ plasma cells showed the presence of all the four IgG subclasses. The results of DNA–DNA checkerboard analysis were in agreement with that of SPR analysis for the detection of P. gingivalis in patient samples. Furthermore, incubation with anti-P. gingivalis attenuated the bacterial response in SPR. The in vitro method for antibody production developed during this study could be used for an efficient real-time detection of periodontitis, and the attenuating effects of in vitro antibodies suggest their role in passive immunization to prevent periodontitis and their associated risk factors.

Place, publisher, year, edition, pages
Wiley-Blackwell, 2015
Keyword
Antibodies, B-cell, plasma cell, periodontitis, P. gingivalis, surface plasmon resonance
National Category
Microbiology in the medical area
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-46728 (URN)10.1002/cre2.6 (DOI)
Available from: 2015-11-24 Created: 2015-11-23 Last updated: 2016-10-06Bibliographically approved
2. Antibacterial effects of lactobacillus and bacteriocin NC8 αβ on the periodontal pathogen Porphyromonas gingivalis
Open this publication in new window or tab >>Antibacterial effects of lactobacillus and bacteriocin NC8 αβ on the periodontal pathogen Porphyromonas gingivalis
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(English)Manuscript (preprint) (Other academic)
National Category
Microbiology in the medical area
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-46730 (URN)
Available from: 2015-11-24 Created: 2015-11-24 Last updated: 2015-11-24Bibliographically approved

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Nakka, Sravya Sowdamini
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