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Louise Chan Bench E 31082016 [1]
Classification
Higher order taxa
Bacteria – Bacteroidetes – Bacteroidia– Bacteroidales – Porphyromonadaceae – Porphyromonas
Species
Species: P. gingivalis
Type Strain: strain 2561 = ATCC 33277= CCUG 25893 = CCUG 25928 = CIP 103683 = DSM 20709 = JCM 12257 = NCTC 11834
Description and significance
Porphyromonas gingivalis was first isolated and identified as Bacteroides melaninogenicus in 1977 by J.Slots at State University of New York, Buffalo [ref]. Slot’s strain 2561 (ATCC 33277) was isolated from a human gingival sulcus during a study of eight patients aged between 34-48 years with advanced periodontitis. In the same year, Finegold and Barnes [ref] refined this classification after their studies revealed that several strains within B. melaninogenicus differed according to their carbohydrate metabolism (asaccharolysis v.s. saccharolysis) and their G + C composition[ref]. In 1980, Coykendall et al. proposed a further reclassification of human oral strains of B. asaccharolyticus to B. gingivalis due to its differences from non oral strains in its base composition, serology and its unique ability to produce phenylacetic acid [ref]. The classification of this microorganism as P. gingivalis did not occur until 1988 when Shah and Collins proposed the reclassification of B. assacharolyticus, B. gingivalis and B. endodontalis in a new genus, Porphyromonas, after the discovery that these three species differed markedly from the type species of Bacteroides, B. fragilis, in many biochemical and chemical properties [ref].
P. gingivalis is a gram negative, non-sporeforming bacteria that forms convex black colonies when cultured on blood agar medium [ref]. These bacteria appear as non-motile long rods or coccobacilli that are approximately 0.5 by 1-2µm in size. It is most commonly found in dental plaque, within the periodontal tissue of patients with periodontitis, and has also occasionally been found in the healthy gingival margin. It has been suggested that due to its presence in the disease and non-disease state, P. gingivalis is an opportunistic pathogen that has co-evolved with Homo sapiens over many years [ref]. A recent study of ancient calcified dental plaque revealed that the abundance of P. gingivalis in dental plaque has been relatively stable over the past 10,000 years. Although P. gingivalis has been identified in other areas of the body, it does not naturally inhabit these regions. Rather, it is through the spread from the oral cavity that these bacteria can be identified in these locations.
Due to the association of P. gingivalis in severe forms of periodontitis, several studies have attempted to identify the mechanism of its pathogenesis. P. gingivalis causes the resorption of alveolar bone at the surface of the tooth root and leads to exfoliation of the teeth. This is mediated through its several virulence factors including lipopolysaccharide (LPS), fimbriae, hemagglutinins, capsular proteins, and proteolytic enzymes. Whilst P. gingivalis possesses virulence factors that contribute to its pathogenesis, it has been suggested that P. gingivalis has an even more significant role of increasing virulence within the community of commensal bacteria. In addition, its ability to evade and subvert the host immune surveillance following the invasion of epithelial cells leads to the problem of persistence of several oral pathogens. P. gingivalis has been termed as a 'keystone' biofilm species that is part of the "Red Complex", a group of bacteria that are categorised by their association with severe forms of periodontitis.
Give a general description of the species (e.g. where/when was it first discovered, where is it commonly found, has it been cultured, functional role, type of bacterium [Gram+/-], morphology, etc.) and explain why it is important to study this microorganism. Examples of citations [1], [2]
Genome structure
Strain ATCC 33277 is the type strain of P. gingivalis. It has a single circular chromosome of 2,354,886 bp and an average G + C content of 48.4%. At this point, no plasmids have been identified in ATCC 33277 [ref]. A large proportion of the genome contains coding sequences. A total of 2090 CDSs have been identified in the genome of ATCC 33277, making up 86.1% of the genome. The genome has 53 tRNA genes that provide the specificity for producing all necessary amino acids and 4 RNA operons.
ATCC 33277 has a total of 93 Insertion Sequence (IS) elements that have the ability to independently transpose, and 48 miniature inverted-repeat transposable elements (MITES) that are dependent on IS element transposase. The genome also has an abundance of other mobile genetic elements including novel conjugative transposons (CTns) encoding Na+-driven multi-drug efflux pumps and composite transposons (Tns) encoding tetR family transcriptional regulators, proteases and ABC transporter ATP-binding proteins. It has previously been proposed that P. gingivalis is assaccharolytic due to a nonsense mutation in the glucose kinase-encoding gene (glk) [ref]. An insertion of MITE239 was identified in the glk gene of ATCC 33277, however, no nonsense mutation was identified. Nonsense mutations were not identified in the glk gene of five other strains of P. gingivalis, suggesting that glk may not be the only determinant of assaccharolysis.
Genes from lateral gene transfer from other bacterial species have been identified. These genes have been proposed to have similarities to genes in other bacteria such as Bacteroides fragilis, the type strain of Bacteroides, and B. thetaiotaomicron, an enteric commensal [ref]. ATCC 33277 is a less virulent strain of P. gingivalis and has been useful in genomic comparative studies for identifying virulence genes in other strains. Many of these virulence genes are located on pathogenicity islands and have been introduced through lateral gene transfer. A comparison of the ATCC 33277 genome to the virulent strain W83 revealed the absence of a cluster of ORFS in ATCC 332277 that are involved in the synthesis of capsular polysaccharide [ref].
Select a strain for which genome information (e.g. size, plasmids, distinct genes, etc.) is available.
Cell structure and metabolism
Cell wall, biofilm formation, motility, metabolic functions.
P. gingivalis is a gram-negative bacteria that contains many virulence factors in its cell wall. Many of these virulence factors such as lipopolysaccharide (LPS), proteases, hemogglutinin, lipoproteins, and fimbriae are found within the outer cell membrane. P. gingivalis LPS mediates a cellular proinflammatory response by activating nuclear factor-κB (NF-κB) and Toll-like receptor 4 (TLR4), inducing the production of cytokines such as interleukin 1 beta (IL1β), interleukin 6 (IL-6) and tumour necrosis factor alpha (TNF-α). P. gingivalis has the ability to utilise more than one innate immune response pathway. Previously it was thought that TLR4 was a distinct receptor in the response to gram-negative bacteria and TLR2 to gram-positive bacteria, however it has now been shown that P. gingivalis has the ability to activate both types of receptors due to the recognition of its multiple lipid A species by the immune cell [ref]. Several proteases are associated with the outer membrane of the cell wall including Arg- and Lys- gingipain cysteine proteinases (Rgp and Kgp), collagenase and aminopeptidases. Aminopeptidase enzymes provide peptides for growth of P. gingivalis. Enzymes such as Gingipains and collagenases neutralise the innate immune system by hydrolysing serum and tissue proteins as well as proenzymes and T-cell receptors [ref]. Hemogglutinin is another protein inbedded within the cell wall. This protein enables P. gingivalis to agglutinate erythrocytes. Hemogglutinin B (HagB) has also been found to have a role in the adhesion of P. gingivalis to host tissue in a similar mechanism to adhesins such as fimbriae. The fimbriae of P. gingivalis are the virulence factors primarily used for adhesion and colonisation. Polymerized fimbrillin (FimA) interacts with extracellular matrix, bacteria and immune cells through modulation by its regulatory protein (FimB) and accessory proteins (FimCDE). Fimbriae facilitate in the invasion of P. gingivalis into host cells through the binding of β1 integrin on the host cell surface. Subsequently, P. gingivalis is actively internalised into the cell and can remain dormant or replicate within epithelial cells without detection by the host immune system. Other proteins that are found within the cell wall ofP. gingivalis include OmpA-like proteins, porins, and lipoproteins such as RagB and RagA that form a receptor complex for the active transportation of hydrolysed proteins.
These several virulence factors facilitate in the colonisation of oral environment and biolfilm formation. P. gingivalis is known as a late coloniser of the periodontal biofilm. Gram positive bacteria such as Streptococci and Actinomyces are early colonisers that initially adhere to the salivary pellicles of the teeth. These are commensal bacteria that are form a niche in the oral cavity.Fusobacterium nucleatum, Treponema denticola and Bacteroides forsythus are the intermediate colonisers that act as bridging bacteria. These bacteria interact with the early colonisers whilst coaggregating with other late colonising microbes such as P. gingivalis. Attachment surfaces are rich in nutrients produced by the early commensal microbiota, signalling molecules that lead to coordinated gene expression, and molecules that in the structural integrity of the biofilm.
Ecology
Aerobe/anaerobe, habitat (location in the oral cavity, potential other environments) and microbe/host interactions.
P. gingivalis is an obligate anaerobe that is most commonly associated with the human oral cavity. Although present at low levels in the healthy human oral microbiome, it is most prevalent within subjects with periodontal disease. As such, its main niche within the oral cavity is on and within the periodontum (tissues damaged by periodontal disease) [ref.]. It has also been shown to survive and replicate within the environmental amoeba Acanthamoeba castellani, suggesting a possible environmental reservoir [ref.].
To aid in its survival within the oral cavity, P. gingivalis can invade the epithelial cells lining the host gingival tissues [ref]. Gingival epithelial cells are typically the first cells to be colonised. These cells are an important first line of defence in the gingival innate immune system. They exhibit the purinergic receptor P2X7, which is responsive to ATP-induced apoptosis [ref.]. However, P. gingivalis has evolved to circumvent this by consuming the ATP signal, thus preventing cell apoptosis [ref.]. Furthermore, it can replicate and disseminate to surrounding cells without rupturing the cell. These features enable P. gingivalis to effectively shield itself from the host immune system.
Pathology
Do these microorganisms cause disease in the oral cavity or elsewhere?
Application to biotechnology
Several different candidate vaccines for P. gingivalis have been developed and theses have focused on using the surface antigens which include capsule, LPS, fimbriae, outer-membrane proteins (OMPs), ginigpains and hemagglutinin [ref.]. Most of these methods have been shown to successfully attenuate the severity of disease. However, the majority of these antigens have several different serotypes and therefore limit the effectiveness of the vaccine to provide complete or adequate protection against all P. gingivalis strains [ref]. Therefore, a change in strategy will be needed to combat this issue. A recent study showed that outer membrane vesicles, which display a wide range of surface antigens, were effective at inhibiting gingipain proteolytic activity in the vaccine strain as well as heterologous strains [1]. Although promising, at present, there has been little if any vaccines trialed with humans and therefore a vaccine against P. ginivalis is still some time off.
Current research
Summarise some of the most recent discoveries regarding this species.
Recent research has highlighted the link between chronic inflammation and systemic disease. Periodontitis is the most common form of chronic inflammation. Association studies have shown that patients suffering from Rheumatoid arthritis have higher rates of clinical attachment loss (CAL), the loss of connective tissue attachment at the periodontium. P. gingivalis is known to invade the epithelial cells lining the gingivae and contribute to this process [Kuboniwa et al.,2008]. Indirect evidence has shown that antibodies targeting P. gingivalis are increased in patients with RA [ref]. In addition, P. gingivalis is unique among oral pathogens for its ability to citrullinate proteins. This is important, as RA is characterised by an autoimmune response to proteins with this modification [ref]. Therefore, this points to periodontal disease and specifically, P. gingivalis, in playing a role in the development of RA.
Periodontal disease is recognised as a risk factor for the development of coronary heart disease, however, the mechanism by which this occurs is still an active field of research. Animal models have shown that disseminated P. gingivalis is capable of causing damage consistent with the histology of cardiovascular disease [ref]. More recent studies are discovering characteristics of P. gingivalis which further elucidate its role in the progession of disease. Liu et al. showed that cholesterol metabolism of macrophages is affected when challenged with P. gingivalis lipopolysaccharide (LPS). Macrophage transformation into lipid-loaded foam cells is an early process in the formation of atherosclerosis (thickening of the arterial walls) [ref]. Another study used a mouse model to show that P. gingivalis enhanced cardiac hypertrophy by increasing the production of reactive oxygen species TLR2 and Nox4 [ref].
An emerging theme in oncology is the role that inflammation, and more specifically, microorganism-induced inflammation, plays in the pathogenesis of cancer. At present, Helicobacter pylori is the only bacteria implicated in the development of cancer, with viruses making up the majority (e.g. Human papillomavirus, Hepatitis B and C etc.). However, the link between inflammation and cancer development is driving more research into the role of organisms such as P. gingivalis due to its association with chronic disease. A recent study by Ha et al. demonstrated that exposing oral squamous cell carcinoma cells to P. gingivalis increased the invasive ability of the oral cancer cells [ref.].
References
References examples
- ↑ MICR3004
This page is written by Louise Chan for the MICR3004 course, Semester 2, 2016
