Fn-OMV potentiates ZBP1-mediated PANoptosis triggered by oncolytic HSV-1 to fuel antitumor immunity.

Oncolytic viruses (OVs) show promise as a cancer treatment by selectively replicating in tumor cells and promoting antitumor immunity. However, the current immunogenicity induced by OVs for tumor treatment is relatively weak, necessitating a thorough investigation of the mechanisms underlying its induction of antitumor immunity. Here, we show that HSV-1-based OVs (oHSVs) trigger ZBP1-mediated PANoptosis (a unique innate immune inflammatory cell death modality), resulting in augmented antitumor immune effects. Mechanistically, oHSV enhances the expression of interferon-stimulated genes, leading to the accumulation of endogenous Z-RNA and subsequent activation of ZBP1. To further enhance the antitumor potential of oHSV, we conduct a screening and identify Fusobacterium nucleatum outer membrane vesicle (Fn-OMV) that can increase the expression of PANoptosis execution proteins. The combination of Fn-OMV and oHSV demonstrates potent antitumor immunogenicity. Taken together, our study provides a deeper understanding of oHSV-induced antitumor immunity, and demonstrates a promising strategy that combines oHSV with Fn-OMV.

Biofilms and core pathogens shape the tumor microenvironment and immune phenotype in colorectal cancer.

Extensive research has explored the role of gut microbiota in colorectal cancer (CRC). Nonetheless, metatranscriptomic studies investigating the in situ functional implications of host-microbe interactions in CRC are scarce. Therefore, we characterized the influence of CRC core pathogens and biofilms on the tumor microenvironment (TME) in 40 CRC, paired normal, and healthy tissue biopsies using fluorescence in situ hybridization (FISH) and dual-RNA sequencing. FISH revealed that Fusobacterium spp. was associated with increased bacterial biomass and inflammatory response in CRC samples. Dual-RNA sequencing demonstrated increased expression of pro-inflammatory cytokines, defensins, matrix-metalloproteases, and immunomodulatory factors in CRC samples with high bacterial activity. In addition, bacterial activity correlated with the infiltration of several immune cell subtypes, including M2 macrophages and regulatory T-cells in CRC samples. Specifically, Bacteroides fragilis and Fusobacterium nucleatum correlated with the infiltration of neutrophils and CD4+ T-cells, respectively. The collective bacterial activity/biomass appeared to exert a more significant influence on the TME than core pathogens, underscoring the intricate interplay between gut microbiota and CRC. These results emphasize how biofilms and core pathogens shape the immune phenotype and TME in CRC while highlighting the need to extend the bacterial scope beyond CRC pathogens to advance our understanding and identify treatment targets.

Gram-negative anaerobes elicit a robust keratinocytes immune response with potential insights into HS pathogenesis.

Hidradenitis Suppurativa (HS) is a chronic autoinflammatory skin disease with activated keratinocytes, tunnel formation and a complex immune infiltrate in tissue. The HS microbiome is polymicrobial with an abundance of commensal gram-positive facultative (GPs) Staphylococcus species and gram-negative anaerobic (GNA) bacteria like Prevotella, Fusobacterium and Porphyromonas with increasing predominance of GNAs with disease severity. We sought to define the keratinocyte response to bacteria commonly isolated from HS lesions to probe pathogenic relationships between HS and the microbiome. Type strains of Prevotella nigrescens, Prevotella melaninogenica, Prevotella intermedia, Prevotella asaccharolytica, Fusobacterium nucleatum, as well as Staphylococcus aureus and the normal skin commensal Staphylococcus epidermidis were heat-killed and co-incubated with normal human keratinocytes. RNA was collected and analysed using RNAseq and RT-qPCR. The supernatant was collected from cell culture for protein quantification. Transcriptomic profiles between HS clinical samples and stimulated keratinocytes were compared. Co-staining of patient HS frozen sections was used to localize bacteria in lesions. A mouse intradermal injection model was used to investigate early immune recruitment. TLR4 and JAK inhibitors were used to investigate mechanistic avenues of bacterial response inhibition. GNAs, especially F. nucleatum, stimulated vastly higher CXCL8, IL17C, CCL20, IL6, TNF and IL36γ transcription in normal skin keratinocytes than the GPs S. epidermidis and S. aureus. Using RNAseq, we found that F. nucleatum (and Prevotella) strongly induced the IL-17 pathway in keratinocytes and overlapped with transcriptome profiles of HS patient clinical samples. Bacteria were juxtaposed to activated keratinocytes in vivo, and F. nucleatum strongly recruited murine neutrophil and macrophage migration. Both the TLR4 and pan-JAK inhibitors reduced cytokine production. Detailed transcriptomic profiling of healthy skin keratinocytes exposed to GNAs prevalent in HS revealed a potent, extensive inflammatory response vastly stronger than GPs. GNAs stimulated HS-relevant genes, including many genes in the IL-17 response pathway, and were significantly associated with HS tissue transcriptomes. The close association of activated keratinocytes with bacteria in HS lesions and innate infiltration in murine skin cemented GNA pathogenic potential. These novel mechanistic insights could drive future targeted therapies.

Fusobacterium nucleatum and oral cancer: a critical review.

There is a growing level of interest in the potential role inflammation has on the initiation and progression of malignancy. Notable examples include Helicobacter pylori-mediated inflammation in gastric cancer and more recently Fusobacterium nucleatum-mediated inflammation in colorectal cancer. Fusobacterium nucleatum is a Gram-negative anaerobic bacterium that was first isolated from the oral cavity and identified as a periodontal pathogen. Biofilms on oral squamous cell carcinomas are enriched with anaerobic periodontal pathogens, including F. nucleatum, which has prompted hypotheses that this bacterium could contribute to oral cancer development. Recent studies have demonstrated that F. nucleatum can promote cancer by several mechanisms; activation of cell proliferation, promotion of cellular invasion, induction of chronic inflammation and immune evasion. This review provides an update on the association between F. nucleatum and oral carcinogenesis, and provides insights into the possible mechanisms underlying it.

Fusobacterium nucleatum enhances the efficacy of PD-L1 blockade in colorectal cancer.

Given that only a subset of patients with colorectal cancer (CRC) benefit from immune checkpoint therapy, efforts are ongoing to identify markers that predict immunotherapeutic response. Increasing evidence suggests that microbes influence the efficacy of cancer therapies. Fusobacterium nucleatum induces different immune responses in CRC with different microsatellite-instability (MSI) statuses. Here, we investigated the effect of F. nucleatum on anti-PD-L1 therapy in CRC. We found that high F. nucleatum levels correlate with improved therapeutic responses to PD-1 blockade in patients with CRC. Additionally, F. nucleatum enhanced the antitumor effects of PD-L1 blockade on CRC in mice and prolonged survival. Combining F. nucleatum supplementation with immunotherapy rescued the therapeutic effects of PD-L1 blockade. Furthermore, F. nucleatum induced PD-L1 expression by activating STING signaling and increased the accumulation of interferon-gamma (IFN-γ)+ CD8+ tumor-infiltrating lymphocytes (TILs) during treatment with PD-L1 blockade, thereby augmenting tumor sensitivity to PD-L1 blockade. Finally, patient-derived organoid models demonstrated that increased F. nucleatum levels correlated with an improved therapeutic response to PD-L1 blockade. These findings suggest that F. nucleatum may modulate immune checkpoint therapy for CRC.

Relationship between Fusobacterium nucleatum and antitumor immunity in colorectal cancer liver metastasis.

Fusobacterium nucleatum has been detected in 8%-13% of human colorectal cancer, and shown to inhibit immune responses against primary colorectal tumors in animal models. Thus, we hypothesized that the presence of F. nucleatum might be associated with reduced T cell density in colorectal cancer liver metastases (CRLM). We quantified F. nucleatum DNA in 181 CRLM specimens using quantitative PCR assay. The densities of CD8+ T cells, CD33+ cells (marker for myeloid-derived suppressor cells [MDSCs]), and CD163+ cells (marker for tumor-associated macrophages [TAMs]) in CRLM tissue were determined by immunohistochemical staining. Fusobacterium nucleatum was detected in eight (4.4%) of 181 CRLM specimens. Compared with F. nucleatum-negative CRLM, F. nucleatum-positive CRLM showed significantly lower density of CD8+ T cells (P = .033) and higher density of MDSCs (P = .001). The association of F. nucleatum with the density of TAMs was not statistically significant (P = .70). The presence of F. nucleatum is associated with a lower density of CD8+ T cells and a higher density of MDSCs in CRLM tissue. Upon validation, our findings could provide insights to develop strategies that involve targeting microbiota and immune cells for the prevention and treatment of CRLM.

Fusobacterium nucleatum and alteration of the oral microbiome: from pregnancy to SARS-COV-2 infection.

OBJECTIVE: The human being has evolved in close symbiosis with its own ecological community of commensal, symbiotic and pathogenic bacteria. After the intestinal microbiome, that of the oral cavity is the largest and most diversified. Its importance is reflected not only in local and systemic diseases, but also in pregnancy since it would seem to influence the placental microbiome. MATERIALS AND METHODS: This is a literature review of articles published in PubMed about Fusobacterium Nucleatum and both its implications with systemic and oral health, adverse pregnancy outcomes, flavors perception and its interference in the oral-nasal mucosal immunity. RESULTS: It is in maintaining the microbiome's homeostasis that the Fusobacterium nucleatum, an opportunistic periodontal pathogen of the oral cavity, plays a crucial role both as a bridge microorganism of the tongue biofilm, and in maintaining the balance between the different species in the oral-nasal mucosal immunity also by taste receptors interaction. It is also involved in the flavor perception and its detection in the oral microbiome of children from the first days of life suggests a possible physiological role. However, the dysbiosis can determine its pathogenicity with local and systemic consequences, including the pathogenesis of respiratory infections. CONCLUSIONS: It is interesting to evaluate its possible correlation with Sars-CoV-2 and the consequences on the microflora of the oral cavity, both to promote a possible broad-spectrum preventive action, in favor of all subjects for whom, by promoting the eubiosis of the oral microbiome, a defensive action could be envisaged by the commensals themselves but, above all, for patients with specific comorbidities and therefore already prone to oral dysbiosis.

Interaction between intestinal microbiota and tumour immunity in the tumour microenvironment.

In recent years, an increasing number of studies have reported that intestinal microbiota have an important effect on tumour immunity by affecting the tumour microenvironment (TME). The intestinal microbiota are closely associated with various immune cells, such as T lymphocytes, natural killer cells (NK cells) and macrophages. Some bacteria, such as Akkermansia muciniphila (A. muciniphila) and Lactobacillus reuteri (L. reuteri), have been shown to improve the effect of tumour immunity. Furthermore, microbial imbalance, such as the increased abundance of Fusobacterium nucleatum (F. nucleatum) and Helicobacter hepaticus (H. hepaticus), generally causes tumour formation and progression. In addition, some microbiota also play important roles in tumour immunotherapy, especially PD-L1-related therapies. Therefore, what is the relationship between these processes and how do they affect each other? In this review, we summarize the interactions and corresponding mechanisms among the intestinal microbiota, immune system and TME to facilitate the research and development of new targeted drugs and provide new approaches to tumour therapy.

Fusobacterium nucleatum Facilitates M2 Macrophage Polarization and Colorectal Carcinoma Progression by Activating TLR4/NF-κB/S100A9 Cascade.

Fusobacterium nucleatum (Fn) has been considered as a significant contributor in promoting colorectal carcinoma (CRC) development by suppressing host anti-tumor immunity. Recent studies demonstrated that the aggregation of M2 macrophage (Mφ) was involved in CRC progress driven by Fn infection. However, the underlying molecular mechanisms are poorly characterized. Here, we investigated the role of Fn in Mφ polarization as well as its effect on CRC malignancy. Fn infection facilitated differentiation of Mφ into the M2-like Mφ phenotype by in vitro study. Histological observation from Fn-positive CRC tissues confirmed the abundance of tumor-infiltrating M2-like Mφ. Fn-induced M2-like Mφ polarization was weakened once inhibiting a highly expressed damage-associated molecular pattern (DAMP) molecule S100A9 mainly derived from Fn-challenged Mφ and CRC cells. In addition, Fn-challenged M2-like Mφ conferred CRC cells a more malignant phenotype, showing stronger proliferation and migration characteristics in vitro and significantly enhanced tumor growth in vivo, all of which were partially inhibited when S100A9 was lost. Mechanistic studies further demonstrated that activation of TLR4/NF-κB signaling pathway mediated Fn-induced S100A9 expression and subsequent M2-like Mφ activation. Collectively, these findings indicate that elevated S100A9 in Fn-infected CRC microenvironment participates in M2-like Mφ polarization, thereby facilitating CRC malignancy. Furthermore, targeting TLR4/NF-κB/S100A9 cascade may serve as promising immunotherapeutic strategy for Fn-associated CRC.

Fusobacterium nucleatum Secretes Outer Membrane Vesicles and Promotes Intestinal Inflammation.

Multiple studies have implicated microbes in the development of inflammation, but the mechanisms remain unknown. Bacteria in the genus Fusobacterium have been identified in the intestinal mucosa of patients with digestive diseases; thus, we hypothesized that Fusobacterium nucleatum promotes intestinal inflammation. The addition of >50 kDa F. nucleatum conditioned media, which contain outer membrane vesicles (OMVs), to colonic epithelial cells stimulated secretion of the proinflammatory cytokines interleukin-8 (IL-8) and tumor necrosis factor (TNF). In addition, purified F. nucleatum OMVs, but not compounds <50 kDa, stimulated IL-8 and TNF production; which was decreased by pharmacological inhibition of Toll-like receptor 4 (TLR4). These effects were linked to downstream effectors p-ERK, p-CREB, and NF-κB. F. nucleatum >50-kDa compounds also stimulated TNF secretion, p-ERK, p-CREB, and NF-κB activation in human colonoid monolayers. In mice harboring a human microbiota, pretreatment with antibiotics and a single oral gavage of F. nucleatum resulted in inflammation. Compared to mice receiving vehicle control, mice treated with F. nucleatum showed disruption of the colonic architecture, with increased immune cell infiltration and depleted mucus layers. Analysis of mucosal gene expression revealed increased levels of proinflammatory cytokines (KC, TNF, IL-6, IFN-γ, and MCP-1) at day 3 and day 5 in F. nucleatum-treated mice compared to controls. These proinflammatory effects were absent in mice who received F. nucleatum without pretreatment with antibiotics, suggesting that an intact microbiome is protective against F. nucleatum-mediated immune responses. These data provide evidence that F. nucleatum promotes proinflammatory signaling cascades in the context of a depleted intestinal microbiome.IMPORTANCE Several studies have identified an increased abundance of Fusobacterium in the intestinal tracts of patients with colon cancer, liver cirrhosis, primary sclerosing cholangitis, gastroesophageal reflux disease, HIV infection, and alcoholism. However, the direct mechanism(s) of action of Fusobacterium on pathophysiological within the gastrointestinal tract is unclear. These studies have identified that F. nucleatum subsp. polymorphum releases outer membrane vesicles which activate TLR4 and NF-κB to stimulate proinflammatory signals in vitro Using mice harboring a human microbiome, we demonstrate that F. nucleatum can promote inflammation, an effect which required antibiotic-mediated alterations in the gut microbiome. Collectively, these results suggest a mechanism by which F. nucleatum may contribute to intestinal inflammation.

Heat-killed Lactobacillus acidophilus mediates Fusobacterium nucleatum induced pro-inflammatory responses in epithelial cells.

Probiotics is widespreadly used nowadays. However, the safety issue with the use of live probiotics is still a matter of contention. In recent years, an expanding body of evidence supports the beneficial role of heat-killed probiotics in the maintenance of systemic health, whereas the role of these heat-killed bacteria on periodontal health remains unclear. This study aimed to evaluate the effects of heat-killed probiotics on periodontal pathogen virulence and associated mechanisms. We demonstrated that heat-killed Lactobacillus acidophilus was able to coaggregate with Fusobacterium nucleatum, the bridging bacteria of oral biofilm, and inhibit the adhesion and invasion of F. nucleatum, leading to a subsequent elimination of pro-inflammatory cytokine production in oral epithelial cells. This coaggregation further caused a suppression of the virulence gene fap2 expression in F. nucleatum. Therefore, heat-killed L. acidophilus might downregulate the pro-inflammatory cytokine expression in epithelial cells via coaggregation with F. nucleatum and suppression of F. nucleatum fap2 expression, which was the first demonstration that heat-killed probiotics modulate periodontal disease pathogenesis via coaggregation. Collectively, this finding provides new evidence that heat-killed probiotics might exert beneficial effects to periodontal health by coaggregating with periodontal pathogens and modulating their virulence.

The onco-immunological implications of Fusobacterium nucleatum in breast cancer.

Breast cancer is a leading cause of death worldwide and a better understanding of this disease is needed to improve treatment outcomes. Recent evidence indicates that bacterial dysbiosis is associated with breast cancer, but the bacteria involved remain poorly characterised. Furthermore, an association between periodontal disease, characterised by oral dysbiosis, and breast cancer have also been discovered, but the mechanisms responsible for this association remains to be elucidated. The oral bacterium involved in periodontal disease, Fusobacterium nucleatum, have recently been detected in human breast tumour tissue and it promoted tumour growth and metastatic progression in a mouse model. The mechanisms of how F. nucleatum might colonise breast tissue and how it might promote tumour progression has not been fully elucidated yet. Here we discuss the breast tumour microbiota, its colonisation by F. nucleatum, possible mechanisms by which F. nucleatum might promote breast cancer progression and how this might impact breast cancer treatment. Literature indicates that F. nucleatum might promote breast cancer progression through activating the Toll-like receptor 4 pathway and by supressing the immune system. This results in cell growth and treatment resistance through autophagy as well as immune evasion. Targeted treatment directed at F. nucleatum combined with immunotherapy and autophagy inhibitors might therefore be a feasible treatment strategy for breast cancer patients.

Prediagnostic Antibody Responses to Fusobacterium nucleatum Proteins Are Not Associated with Risk of Colorectal Cancer in a Large U.S. Consortium.

BACKGROUND: The association between prediagnostic antibody responses to Fusobacterium nucleatum (F. nucleatum) and subsequent risk of colorectal cancer is not established. METHODS: We conducted a nested case-control study of 8,126 participants in a consortium of 10 prospective cohorts in the United States. RESULTS: Higher seroprevalence of any F. nucleatum antibody was observed among non-White participants (51.1%) compared with White participants (31.2%). We did not find any statistically significant association between seropositivity to any of the eight F. nucleatum proteins and colorectal cancer risk. CONCLUSIONS: Prediagnostic antibody responses to F. nucleatum proteins were not associated with the risk of colorectal cancer. IMPACT: Future studies may consider a more specific detection of the immunoglobulin isotypes or focus on examining F. nucleatum in stool or tissue samples.

Diagnostic accuracy of Fusobacterium nucleatum IgA and IgG ELISA test in colorectal cancer.

The colorectal cancer is a serious health problem. The diagnosis of the disease mostly relies on an invasive procedure. A non-invasive diagnostic test such as an immunoassay, may facilitate diagnosis of colorectal cancer. The purpose of the study was to evaluate the use of antibodies against Fusobacterium nucleatum in the diagnosis of colorectal cancer (CRC). Totally 78 patients in three groups were included in the study. F. nucleatum in the tissues was detected using quantitative polymerase chain reaction assay. F. nucleatum IgA and IgG were measured using enzyme linked immunosorbent assay. F. nucleatum was detected in 86.7% and 73.1% cases of CRC and precancerous-benign colon disease (P-BCD), respectively. The OD values from F. nucleatum IgA and IgG ELISA tests were higher in CRC group compared with healthy individuals. The sensitivity of IgA ELISA test varied between 31.8 and 95.5% depending on the chosen cut-off values. The positivity rate of antibodies in patients with high amount of F. nucleatum in tissue was significantly greater than in the negative group. The F. nucleatum IgA and IgG antibodies in CRC were higher than the ones in healthy controls but the discriminative ability of the ELISA test was not adequate to be considered as a diagnostic tool.

Fusobacteriumnucleatum Adheres to Clostridioides difficile via the RadD Adhesin to Enhance Biofilm Formation in Intestinal Mucus.

BACKGROUND & AIMS: Although Clostridioides difficile infection (CDI) is known to involve the disruption of the gut microbiota, little is understood regarding how mucus-associated microbes interact with C difficile. We hypothesized that select mucus-associated bacteria would promote C difficile colonization and biofilm formation. METHODS: To create a model of the human intestinal mucus layer and gut microbiota, we used bioreactors inoculated with healthy human feces, treated with clindamycin and infected with C difficile with the addition of human MUC2-coated coverslips. RESULTS: C difficile was found to colonize and form biofilms on MUC2-coated coverslips, and 16S rRNA sequencing showed a unique biofilm profile with substantial cocolonization with Fusobacterium species. Consistent with our bioreactor data, publicly available data sets and patient stool samples showed that a subset of patients with C difficile infection harbored high levels of Fusobacterium species. We observed colocalization of C difficile and F nucleatum in an aggregation assay using adult patients and stool of pediatric patients with inflammatory bowel disease and in tissue sections of patients with CDI. C difficile strains were found to coaggregate with F nucleatum subspecies in vitro; an effect that was inhibited by blocking or mutating the adhesin RadD on Fusobacterium and removal of flagella on C difficile. Aggregation was shown to be unique between F nucleatum and C difficile, because other gut commensals did not aggregate with C difficile. Addition of F nucleatum also enhanced C difficile biofilm formation and extracellular polysaccharide production. CONCLUSIONS: Collectively, these data show a unique interaction of between pathogenic C difficile and F nucleatum in the intestinal mucus layer.

Oral microbial dysbiosis and its performance in predicting oral cancer.

Dysbiosis of oral microbiome may dictate the progression of oral squamous cell carcinoma (OSCC). Yet, the composition of oral microbiome fluctuates by saliva and distinct sites of oral cavity and is affected by risky behaviors (smoking, drinking and betel quid chewing) and individuals' oral health condition. To characterize the disturbances in the oral microbial population mainly due to oral tumorigenicity, we profiled the bacteria within the surface of OSCC lesion and its contralateral normal tissue from discovery (n = 74) and validation (n = 42) cohorts of male patients with cancers of the buccal mucosa. Significant alterations in the bacterial diversity and relative abundance of specific oral microbiota (most profoundly, an enrichment for genus Fusobacterium and the loss of genus Streptococcus in the tumor sites) were identified. Functional prediction of oral microbiome shown that microbial genes related to the metabolism of terpenoids and polyketides were differentially enriched between the control and tumor groups, indicating a functional role of oral microbiome in formulating a tumor microenvironment via attenuated biosynthesis of secondary metabolites with anti-cancer effects. Furthermore, the vast majority of microbial signatures detected in the discovery cohort was generalized well to the independent validation cohort, and the clinical validity of these OSCC-associated microbes was observed and successfully replicated. Overall, our analyses reveal signatures (a profusion of Fusobacterium nucleatum CTI-2 and a decrease in Streptococcus pneumoniae) and functions (decreased production of tumor-suppressive metabolites) of oral microbiota related to oral cancer.

Differential immune microenvironmental features of microsatellite-unstable colorectal cancers according to Fusobacterium nucleatum status.

It has been suggested that Fusobacterium nucleatum (Fn) may differentially impact tumor immune responses according to microsatellite instability (MSI) status in colorectal cancers (CRCs). We aimed to reveal the detailed relationship between intratumoral Fn and immune microenvironmental features in MSI-high CRCs. A total of 126 MSI-high CRCs were subjected to analyses for intratumoral Fn DNA load using quantitative PCR and for densities of tumor-infiltrating immune cells, including CD3+ T cells, CD4+ T cells, CD8+ T cells, FoxP3+ T cells, CD68+ macrophages, CD163+ macrophages, and CD177+ neutrophils, at invasive margin (IM) and center of tumor (CT) areas using computational image analysis of immunohistochemistry. Based on the Fn load, the 126 MSI-high CRCs were classified into Fn-high, -low, and -negative subgroups. The Fn-high subset of MSI-high CRCs was significantly correlated with larger tumor size and advanced invasion depth (p = 0.017 and p = 0.034, respectively). Compared with the Fn-low/negative subgroup, Fn-high tumors demonstrated significantly lower density of FoxP3+ cells in both IM and CT areas (p = 0.002 and p = 0.003, respectively). Additionally, Fn-high was significantly associated with elevated CD163+ cell to CD68+ cell ratio in only CT areas of MSI-high CRCs (p = 0.028). In conclusion, the Fn-enriched subset of MSI-high CRCs is characterized by increased tumor growth and invasion and distinct immune microenvironmental features, including decreased FoxP3+ T cells throughout the tumor and increased proportion of M2-polarized macrophages in the tumor center. These findings collectively support that Fn may be linked to pro-tumoral immune responses in MSI-high CRCs.

Identification of Proteins Associated with the Formation of Oral Biofilms

ABSTRACT Objective: To identify proteins associated with the formation of Streptococcus gordonii and Fusobacterium nucleatum biofilms. Material and Methods: Biofilms composed of two bacterial species, S. gordonii and F. nucleatum, were cultured for 1, 4, 7, and 10 days. The presence of both species was confirmed via amplification of the srtA and radD genes using real-time PCR. The concentrations of proteins associated with the biofilms and individual species were quantified using Western blotting. Results: The protein profiles of S. gordonii and F. nucleatum from individual cultures determined using one-dimensional electrophoresis revealed proteins found in S. gordonii and in F. nucleatum. Ct and reciprocal Ct values were determined for the exposed S. gordonii and F. nucleatum biofilms. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) protein was detected in biofilms and F. nucleatum, whereas HSP40 protein was present only in biofilms after 7 and 10 days of formation. Conclusion: HSP40 was detected only in the formed biofilms; thus, HSP40 is an essential proteins for adhesion.

Structural analysis of the lipopolysaccharide O-antigen from Fusobacterium nucleatum strain CC 7/3 JVN3 C1 and development of a mouse monoclonal antibody specific to the O-antigen.

Fusobacterium nucleatum is becoming increasingly recognised as an emerging pathogen, gaining attention as a potential factor for exacerbating colorectal cancer and is strongly linked with pregnancy complications including pre-term and still births. Little is known about the virulence factors of this organism; thus, we have initiated studies to examine the bacterium's surface glycochemistry. In an effort to characterise the surface carbohydrates of F. nucleatum, the aims of this study were to investigate the structure of the lipopolysaccharide (LPS) O-antigen of the cancer-associated isolate F. nucleatum strain CC 7/3 JVN3 C1 (hereafter C1) and to develop monoclonal antibodies (mAbs) to the LPS O-antigen that may be beneficial to the growing field of F. nucleatum research. In this study, we combined several technologies, including nuclear magnetic resonance (NMR) spectroscopy, to elucidate the structure of the LPS O-antigen repeat unit as -[-4-ß-Gal-3-α-FucNAc4N-4-α-NeuNAc-]-. We have previously identified this structure as the LPS O-antigen repeat unit from strain 10953. In this present study, we developed a mAb to the C1 LPS O-antigen and confirmed the mAbs cross-reactivity to the 10953 strain, thus confirming the structural identity.

Serum immunoglobulin G antibody titer to Fusobacterium nucleatum is associated with unfavorable outcome after stroke.

Stroke can be a cause of death, while in non-fatal cases it is a common cause of various disabilities resulting from associated brain damage. However, whether a specific periodontal pathogen is associated with increased risk of unfavorable outcome after stroke remains unknown. We examined risk factors for unfavorable outcome following stroke occurrence, including serum antibody titers to periodontal pathogens. The enrolled cohort included 534 patients who had experienced an acute stroke, who were divided into favorable (n = 337) and unfavorable (n = 197) outcome groups according to modified ranking scale (mRS) score determined at 3 months after onset (favorable = score 0 or 1; unfavorable = score 2-6). The associations of risk factors with unfavorable outcome, including serum titers of IgG antibodies to 16 periodontal pathogens, were examined. Logistic regression analysis showed that the initial National Institutes of Health stroke scale score [odds ratio (OR) = 1·24, 95% confidence interval (CI) = 1·18-1·31, P < 0·001] and C-reactive protein (OR = 1·29, 95% CI = 1·10-1·51, P = 0·002) were independently associated with unfavorable outcome after stroke. Following adjustment with those, detection of the antibody for Fusobacterium nucleatum ATCC 10953 in serum remained an independent predictor of unfavorable outcome (OR = 3·12, 95% CI = 1·55-6·29, P = 0·002). Determination of the antibody titer to F. nucleatum ATCC 10953 in serum may be useful as a predictor of unfavorable outcome after stroke.