Given the potential relationship between head and neck squamous cell carcinoma (HNSCC) and microbial dysbiosis, we profiled the microbiome within healthy normal and tumorous (primary and metastatic) human tissues from the oral cavity, larynx-pharynx, and lymph nodes using 16S rRNA sequencing. Alpha and beta diversity analyses revealed that normal tissues had the greatest richness in community diversity, while the metastatic populations were most closely related to one another. Compared to the normal, the microbiota associated with tumors supported altered abundances in the phyla Fusobacteria, Firmicutes, Actinobacteria and Proteobacteria. Most notably, the relative abundance of Fusobacterium increased whereas Streptococcus decreased in both primary and metastatic samples. Principal coordinate analysis indicated a separation and clustering of samples by tissue status. However, random forest analysis revealed that the microbial profiles alone were a poor predictor for primary and metastatic HNSCC samples. Here, we report that the microbial communities residing in the tumorous tissues are compositionally distinct compared to the normal adjacent tissues. However, likely due to the smaller sample size and sample-to-sample heterogeneity, our prediction models were not able to distinguish by sample types. This work provides a foundation for future studies aimed at understanding the role of the dysbiotic tissue microbiome in HNSCC.

Oral pathogens, including Treponema denticola, initiate the dysregulation of tissue homeostasis that characterizes periodontitis. However, progress of research on the roles of T. denticola in microbe-host interactions and signaling, microbial communities, microbial physiology, and molecular evolution has been hampered by limitations in genetic methodologies. This is typified by an extremely low transformation efficiency and inability to transform the most widely studied T. denticola strain with shuttle plasmids. Previous studies have suggested that robust restriction-modification (R-M) systems in T. denticola contributed to these problems. To facilitate further molecular genetic analysis of T. denticola behavior, we optimized existing protocols such that shuttle plasmid transformation efficiency was increased by >100-fold over prior reports. Here, we report routine transformation of T. denticola ATCC 35405 with shuttle plasmids, independently of both plasmid methylation status and activity of the type II restriction endonuclease encoded by TDE0911. To validate the utility of this methodological advance, we demonstrated expression and activity in T. denticola of a flavin mononucleotide-based fluorescent protein (FbFP) that is active under anoxic conditions. Addition of routine plasmid-based fluorescence labeling to the Treponema toolset will enable more-rigorous and -detailed studies of the behavior of this organism.

Introduction

Nisin, a broad-spectrum bacteriocin, has recently been highlighted for its biomedical applications. To date, no studies have examined the antimicrobial and antibiofilm properties of high-purity (>95%) nisin (nisin ZP) on Enterococcus faecalis and biofilms formed by this species. We hypothesize that nisin can inhibit E. faecalis and reduce biofilm biomass, and combinations of nisin and sodium hypochlorite (NaOCl) will enhance the antibiofilm properties against E. faecalis biofilms.

Methods

Using broth cultures, disc diffusion assays, and biofilm assays, we examined the effects of nisin on various E. faecalis growth parameters and biofilm properties (biovolume, thickness, and roughness). Confocal microscopy was used in conjunction with Imaris and Comstat2 software (Kongens Lyngby, Copenhagen, Denmark) to measure and analyze the biofilm properties.

Results

Nisin significantly decreased the growth of planktonic E. faecalis dose dependently. The minimum inhibitory concentrations against E. faecalis strains OG-1 and ATCC 29212 were 15 and 50 μg/mL, and the minimum bactericidal concentrations were 150 and 200 μg/mL, respectively. A reduction in biofilm biovolume and thickness was observed for biofilms treated with nisin at ≥10 μg/mL for 10 minutes. In addition, the combination of nisin with low doses of NaOCl enhanced the antibiofilm properties of both antimicrobial agents.

Conclusions

Nisin alone or in combination with low concentrations of NaOCl reduces the planktonic growth of E. faecalis and disrupts E. faecalis biofilm structure. Our results suggest that nisin has potential as an adjunctive endodontic therapeutic agent and as an alternative to conventional NaOCl irrigation.

Periodontitis has been associated with many systemic diseases and conditions, including metabolic syndrome. Metabolic syndrome is a cluster of conditions that occur concomitantly and together they increase the risk of cardiovascular disease and double the risk of type 2 diabetes. In this review, we focus on the association between metabolic syndrome and periodontitis; however, we also include information on diabetes mellitus and cardiovascular disease, since these two conditions are significantly intertwined with metabolic syndrome. With regard to periodontitis and metabolic syndrome, to date, the vast majority of studies point to an association between these two conditions and also demonstrate that periodontitis can contribute to the development of, or can worsen, metabolic syndrome. Evaluating the effect of metabolic syndrome on the salivary microbiome, data presented herein support the hypothesis that the salivary bacterial profile is altered in metabolic syndrome patients compared with healthy patients. Considering periodontitis and these three conditions, the vast majority of human and animal studies point to an association between periodontitis and metabolic syndrome, diabetes, and cardiovascular disease. Moreover, there is evidence to suggest that metabolic syndrome and diabetes can alter the oral microbiome. However, more studies are needed to fully understand the influence these conditions have on each other.