ABSTRACT: Dental abscess in ruminants is an acute polymicrobial infection, usually resulting from periodontal disease or endodontic infection, with consequences for animal health and welfare. The present study aimed to describe the bacterial microbiota of dental abscesses in Blastocerus dichotomus. Biological material from mandibular dental abscesses, punctured with a sterile syringe and needle during routine veterinary curative procedures or necropsies, was collected from three ex-situ marsh deer. Bacteria were identified using high-throughput sequencing of the 16S ribosomal RNA gene. The three specimens had the presence of facial bulging, and two died because of severe emaciation with a history of progressive weight loss. Bacteroides (38.6%), Fusobacterium (36.65%), and Porphyromonas (7.49%) represented the most abundant genera and Fusobacterium necrophorum (35.69%), Porphyromonas levii (3.12%) and Porphyromonas gulae (1.78%) were among the ten most represented species in the microbiota of mandibular abscess in Blastocerus dichotomus. These molecular findings demonstrate a broader diversity of species in the polymicrobial nature of dental abscesses in B. dichotomus than was previously reported when culture-dependent methods were used in the diagnosis.

Feline odontoclastic resorptive lesion (FORL) is an inflammatory oral disease of unknown aetiopathogenesis that affects between 20% to 75% of cats. Twenty immune-associated molecules were measured in saliva of 25 healthy and 40 cats with FORL using a multiplex assay. No statistically significant differences were observed in the levels of these proteins between the healthy group and the diseased group of cats. A two-step cluster analysis of the oral microbiome and salivary cytokine data identified two subgroups of cats with FORL: FORL-1 (subset of cats with a less diverse oral microbiome) and FORL-2 (diseased cats with a microbiome similar to that of healthy animals). The level of some key proinflammatory cytokines (IL-1β, IL-12p40) and chemokines (IL-8, RANTES, KC) were significantly higher in the FORL-1 subgroup than in the FORL-2 subgroup and the healthy group. In addition, TNF-α levels were greater in the FORL-1 subgroup than in the FORL-2 subgroup. These increases in pro-inflammatory cytokines and chemokines indicate active ongoing inflammation that may promote the osteoclastic/odontoclastic activity associated with FORL.

The global clinical and socioeconomic impact of chronic wounds is substantial. The main difficulty that clinicians face during the treatment of chronic wounds is the risk of infection at the wound site. Infected wounds arise from an accumulation of microbial aggregates in the wound bed, leading to the formation of polymicrobial biofilms that can be largely resistant to antibiotic therapy. Therefore, it is essential for studies to identify novel therapeutics to alleviate biofilm infections. One innovative technique is the use of cold atmospheric plasma (CAP) which has been shown to possess promising antimicrobial and immunomodulatory properties. Here, different clinically relevant biofilm models will be treated with cold atmospheric plasma to assess its efficacy and killing effects. Biofilm viability was assessed using live dead qPCR, and morphological changes associated with CAP evaluated using scanning electron microscopy (SEM). Results indicated that CAP was effective against Candida albicans and Pseudomonas aeruginosa, both as mono-species biofilms and when grown in a triadic model system. CAP also significantly reduced viability in the nosocomial pathogen, Candida auris. Staphylococcus aureus Newman exhibited a level of tolerance to CAP therapy, both when grown alone or in the triadic model when grown alongside C. albicans and P. aeruginosa. However, this degree of tolerance exhibited by S. aureus was strain dependent. At a microscopic level, biofilm treatment led to subtle changes in morphology in the susceptible biofilms, with evidence of cellular deflation and shrinkage. Taken together, these results indicate a promising application of direct CAP therapy in combatting wound and skin-related biofilm infections, although biofilm composition may affect the treatment efficacy.

Extensive cattle livestock is advancing in Amazonia and its low productivity, with consequent pressure to open new areas, is partly due to sanitary problems and, among them, the periodontal diseases, whose environmental triggers or modifying factors are unknown. In this study, we used high-throughput sequencing, network analysis and predicted functions to investigate the dental and ruminal microbiota of cattle raised in new livestock areas in the Amazon and identify possible keystone pathogens and proteins associated with the disease. Ninety-three genera were common in dental and ruminal fluid microbiomes and among them periodontal pathogens such as Fusobacterium, Prevotella, Porphyromonas and Actinomyces were recognized. Network analysis showed that dental microbiomes of clinically healthy animals tend to comprise a group of OTUs in homeostasis and when analyzed together, dental and ruminal fluid microbiomes of animals with periodontitis had almost twice the number of negative edges, indicating possible competition between bacteria and dysbiosis. The incisor dental and ruminal fluid microbiomes were dominated by a core community composed of members of the phyla Firmicutes and Bacteroidetes. Network results showed that members of the Prevotella genus stood out among the top five OTUs, with the largest number of hubs in the dental and ruminal microbiota of animals with periodontitis. Protein families linked to an inflammatory environment were predicted in the dental and ruminal microbiota of cattle with periodontitis. The dissimilarity between dental microbiomes, discriminating between healthy cattle and those with periodontitis and the identification of possible key pathogens, represent an important reference to elucidate the triggers involved in the etiopathogenesis of bovine periodontitis, and possibly in the development of measures to control the disease and reduce the pressures for deforestation.

Periodontitis is a polymicrobial biofilm-induced inflammatory disease associated with a dysbiotic microbial community and severely affects the health and welfare of animals. However, little is known regarding the dental microbiota associated with this disease in goats. In this study, we used high-throughput sequencing, network analysis, and predicted functions to investigate the microbiota of clinically healthy goats and those with periodontitis and identify possible pathogens and proteins associated with the disease. Dental microbiomes of goats with periodontitis were richer, and network analyses showed that the number of negative interactions was higher in the networks of animals with periodontitis. Based on the interrelationships, Porphyromonas, Fusobacterium, and Prevotella were suggested to play an important role in the dental microbiota associated with goat periodontitis. Protein families linked to translation, cytoplasmatic translation, and rRNA processing were more abundant in the dental microbiota of goats with periodontitis. In conclusion, the dental biofilm microbiota associated with goat periodontitis seems to be dysbiotic and has significant antagonistic interactions, which discriminate healthy animals from diseased animals and highlight the importance of key bacteria. Thus, these novel findings contribute to the evolution of knowledge regarding the etiopathogenesis of goat periodontitis and possibly to the development of periodontitis control measures.

Feline odontoclastic resorptive lesion (FORL) is a common chronic inflammatory condition whose aetiopatho-genesis remains unclear. FORL affects 20-75% of cats and causes excruciating pain and tooth loss. The purpose of this study was to evaluate chronic inflammation in FORL by assessing differences in Toll-like receptor (TLR) and cytokine transcripts in gingival tissues between diseased and healthy cats. Gingival tissue samples were collected from 14 healthy cats with no known clinical signs of oral disease and 41 cats with FORL. Levels of mRNA encoding TLR2, TLR3, TLR4, TLR7, TLR9 and the cytokines interleukin-1 beta (IL-1 beta), IL-4, IL-6, IL-10, IL-12, interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) was evaluated using quantitative real-time PCR. Statistical significance of the results was assessed using non-parametric tests. Levels of TLR and cytokine transcripts were upregulated in gingival tissue from cats with FORL as compared with healthy gingival tissue: TLR2, TLR3 and TLR9, p <= 0.001; TLR4 and TLR7, p <= 0.01; IFN-gamma, IL-4, IL-6, IL-10, IL-12, IL-1 beta and TNF-alpha, p <= 0.001). In conclusion, expression of TLR and both pro-and anti-inflammatory cytokines were significantly increased, confirming an ongoing chronic inflammatory response to the microbiome in FORL. It is likely that dysbiosis of the oral microbiota in cats with FORL activates the innate immune response, leading to active inflammation that results in tooth resorption.

In the mid-1960s the microbial aetiology of periodontal diseases was introduced based on classical experimental gingivitis studies . Since then, numerous studies have addressed the fundamental role that oral microbiota plays in the initiation and progression of periodontal diseases. Recent advances in laboratory identification techniques have contributed to a better understanding of the complexity of the oral microbiome in both health and disease. Modern culture-independent methods such as human oral microbial identification microarray and next-generation sequencing have been used to identify a wide variety of microbial taxa residing in the gingival sulcus and the periodontal pocket. The first theory of the ‘non-specific plaque’ hypothesis gave rise to the ‘ecological plaque’ hypothesis and more recently to the ‘polymicrobial synergy and dysbiosis hypothesis’. Periodontitis is now considered to be a multimicrobial inflammatory disease in which the various bacterial species within the dental biofilm are in a dysbiotic state and this imbalance favours the establishment of chronic inflammatory conditions and ultimately the destruction of tooth-supporting tissues. Apart from the known putative periodontal pathogens, the whole biofilm community is now considered to play a role in the establishment of inflammation and the initiation and progression of periodontitis in a susceptible host. Treatment is unlikely to eliminate putative pathogens but, when it is thoroughly performed it has the potential to establish a healthy ecosystem by altering the microbial community in numbers and composition and also contribute to the maturation of the host immune response.

The work presented in this pilot study aimed to identify potential risk factors associated with bovine periodontitis development. Bovine periodontitis is a multifactorial polymicrobial infectious disease for which the aetiopathogenesis and risk factors are not fully understood. From cattle slaughtered in an abattoir in Scotland, 35 dental arcades with periodontal lesions and 40 periodontally healthy arcades were selected over seven visits for study. Multivariable logistic regression analysis was used to evaluate the association between periodontitis and the independent variables, gender, age and breed. For every increase in year of age, cattle were 1.5 times more likely to have periodontitis. A graphical analysis indicated that within the limits of this study, we could not detect any major influence of breed on the age-effect. Although logistic regression analysis demonstrated that periodontitis lesions are more prevalent with increasing age of cattle the underlying mechanisms remain unclear. It is likely that periodontitis is an important cause of oral pain in older cattle and can contribute to reduced productivity/performance. Further studies with a larger sample size are necessary to elucidate the associations between potential risk factors and periodontitis in cattle and to define its effects on animal welfare and productivity.

Introduction. Periodontitis, one of the most common oral disorders in sheep, is caused by a mixed and opportunistic microbiota that severely affects the health and welfare of animals. However, little is known about the ecological processes involved and the composition of the microbiota associated with the development of the disease. Hypothesis/Gap Statement. Using high-throughput sequencing of the 16S ribosomal RNA gene and network analysis it would be possible to discriminate the microbiomes of clinically healthy sheep and those with periodontitis and possibly identify the key microorganisms associated with the disease. Aim. The present study aimed to characterise the composition of dental microbiomes and bacterial co-occurrence networks in clinically healthy sheep and animals with periodontitis. Methodology. Dental biofilm samples were collected from ten sheep with periodontitis and ten clinically healthy animals. Bacteria were identified using high-throughput sequencing of the 16S ribosomal RNA gene. Results. The most prevalent genera in the dental microbiota of sheep with periodontitis were Petrimonas , Acinetobacter , Porphyromonas and Aerococcus . In clinically healthy animals, the most significant genera were unclassified Pasteurellaceae, Pseudomonas, and Neisseria. Fusobacterium was found at high prevalence in the microbiomes of both groups. The dental microbiota of sheep in the two clinical conditions presented different profiles and the diversity and richness of bacteria was greater in the diseased animals. Network analyses showed the presence of a large number of antagonistic interactions between bacteria in the dental microbiota of animals with periodontitis, indicating the occurrence of a dysbiotic community. Through the interrelationships, members of the Prevotella genus are likely to be key pathogens, both in the dental microbiota of healthy animals and those with periodontitis. Porphyromonas stood out among the top three nodes with more centrality and the largest number of hubs in the networks of animals with periodontitis. Conclusion. The dental biofilm microbiota associated with ovine periodontitis is dysbiotic and with significant antagonistic interactions, which discriminates healthy animals from diseased animals and highlights the importance of key bacteria, such as Petrimonas , Porphyromonas , Prevotella and Fusobacterium species.

Introduction. Feline odontoclastic resorptive lesion (FORL) is one of the most common and painful oral diseases of the cat. It is characterised by tooth resorption due to destructive activity of odontoclasts. FORL can result in tooth loss. While the aetiology of FORL is not clearly understood, it is thought to be multifactorial and bacteria are likely to play a major role. Hypothesis. Dysbiosis of the normal feline oral microbiota leads to an alteration in commensal bacteria populations, which results in the development of FORL. Aim. The purpose of the current study was to determine the composition of the microbiomes associated with feline oral health and FORL. Methodology. Supragingival plaque was collected from 25 cats with a healthy oral cavity and 40 cats with FORL. DNA was extracted from each sample, the V4 region of the 16S rRNA gene amplified by polymerase chain reaction and amplicons sequenced. Diversity and species richness analyses were performed, principal component analysis was used to explore differences between the oral microbiomes of healthy cats and those with FORL, and linear discriminant analysis effect size was used to assess differences between the groups. Results. The six most abundant bacterial genera identified were Bergeyella , Capnocytophaga, Lampropedia , Morexella , Porphyromonas and Treponema . Two-step cluster analysis of the data identified two FORL sub-groups (FORL-1, FORL-2). The FORL-2 sub-group was very similar to the healthy group, whilst the FORL-1 sub-group was clearly different from both the FORL-2 sub-group and the healthy groups. In this analysis, Capnocytophaga ( P <0.001) and Lampropedia ( P <0.01) were found at significantly lower levels and Porphyromonas at a slightly higher level in the FORL-1 sub-group compared to the healthy and FORL-2 sub-groups. Microbial diversity was found to be less in the FORL-1 sub-group than in the healthy group. Lampropedia sp., a phosphate-accumulating oral commensal species, was significantly lower in the FORL-1 sub-group. Conclusion. The oral microbiota associated with the FORL-1 sub-group is distinct from that found in the healthy group and FORL-2 sub-group. Lampropedia species may influence the local calcium-phosphate ratio, which could be a factor in tooth and bone resorption observed in FORL.

INTRODUCTION:The biomedical sciences (BMS) are a central part of the dental curriculum that underpins teaching and clinical practice in all areas of dentistry. Although some specialist groups have proposed curricula in their particular topic areas, there is currently no overarching view of what should be included in a BMS curriculum for undergraduate dental programmes. To address this, the Association for Dental Education in Europe (ADEE) convened a Special Interest Group (SIG) with representatives from across Europe to develop a consensus BMS curriculum for dental programmes.CURRICULUM:This paper summarises the outcome of the deliberations of this SIG and details a consensus view from the SIG of what a BMS curriculum should include.CONCLUSIONS:Given the broad nature of BMS applied to dentistry, this curriculum framework is advisory and seeks to provide programme planners with an indicative list of topics which can be mapped to specific learning objectives within their own curricula. As dentistry becomes increasingly specialised, these will change, or some elements of the undergraduate curriculum may move to the post-graduate setting. So, this document should be seen as a beginning and it will need regular review as BMS curricula in dentistry evolve.

Over the past century, numerous studies have used oral biofilm models to investigate growth kinetics, biofilm formation, structure and composition, antimicrobial susceptibility and host-pathogen interactions. In vivo animal models provide useful models of some oral diseases; however, these are expensive and carry vast ethical implications. Oral biofilms grown or maintained in vitro offer a useful platform for certain studies and have the advantages of being inexpensive to establish and easy to reproduce and manipulate. In addition, a wide range of variables can be monitored and adjusted to mimic the dynamic environmental changes at different sites in the oral cavity, such as pH, temperature, salivary and gingival crevicular fluid flow rates, or microbial composition. This review provides a detailed insight for early-career oral science researchers into how the biofilm models used in oral research have progressed and improved over the years, their advantages and disadvantages, and how such systems have contributed to our current understanding of oral disease pathogenesis and aetiology.

Bovine periodontitis is a progressive and purulent infection associated with an anaerobic subgingival biofilm, which induces irreversible damage to the dentition of affected animals. The aetiopathogenesis of the disease is unclear and treatment and control of the disease process in cattle are almost unknown. The aim of this study was to investigate the innate immune response by quantifying expression of Toll-like receptor (TLR) and cytokine genes in gingival tissue samples from cattle with and without periodontitis. Postmortem biopsies of gingival tissues were collected from 20 cattle with periodontitis and 20 cattle with no clinical signs of periodontal lesions. Tissue expression of TLR2, TLR4, TNF-α, IFN-γ, IL-1β and IL-4 genes were determined using quantitative real-time PCR. Statistically significant increases in mRNA levels encoding TLR2 (p = 0.025), TLR4 (p = 0.037), TNF-α (p = 0.025), IFN-γ (p = 0.014), IL-1β (p < 0.001) and IL-4 (p = 0.014) were observed in animals with periodontitis when compared to periodontally healthy animals. Increased levels of TLRs and inflammatory cytokines in periodontal tissue indicate an induction of the innate immune response of cattle and suggest that a substantial microbial challenge may be involved in the aetiopathogenesis of bovine periodontitis.

Periodontitis is an infectious polymicrobial, immuno-inflammatory disease of multifactorial aetiology that has an impact on the health, production and welfare of ruminants. The objective of the present study was to determine the microbial profiles present in the gingival sulcus of cattle considered periodontally healthy and in the periodontal pocket of animals with periodontitis lesions using high-throughput bacterial 16S rRNA gene sequencing. Subgingival biofilm samples were collected from 40 cattle with periodontitis and 38 periodontally healthy animals. In total, 1923 OTUs were identified and classified into 395 genera or higher taxa. Microbial profiles in health differed significantly from periodontitis in their composition (p < 0.0001, F = 5.30; PERMANOVA) but no statistically significant differences were observed in the diversity of healthy and periodontitis microbiomes. The most prevalent taxa in health were Pseudomonas, Burkholderia and Actinobacteria, whereas in disease these were Prevotella, Fusobacterium and Porphyromonas. The most discriminative taxa in health were Gastranaerophilales, Planifilum and Burkholderia, and in disease these were Elusimicrobia, Synergistes and Propionivibrio. In conclusion, statistically significant difference exists between the microbiome in bovine oral health and periodontitis, with populations showing 72.6% dissimilarity. The diversity of the bacteria found in health and periodontitis were similar and bacteria recognised as periodontal pathogens showed increased abundance in disease. In this context, the main components of bacterial homeostasis in the biofilm of healthy sites and of dysbiosis in periodontal lesions provide unprecedented indicators for the evolution of knowledge about bovine periodontitis.

Periodontitis is a polymicrobial infectious disease that causes occlusion change, tooth loss, difficulty in rumination, and premature culling of animals. This study aimed to detect species of the genera Porphyromonas and Prevotella present in the periodontal pocket of sheep with lesions deeper than 5mm (n=14) and in the gingival sulcus of animals considered periodontally healthy (n=20). The presence of microorganisms was evaluated by polymerase chain reaction (PCR) using specific primers for Porphyromonas asaccharolytica, Porphyromonas endodontalis, Porphyromonas gingivalis, Porphyromonas gulae, Prevotella buccae, Prevotella intermedia, Prevotella loescheii, Prevotella melaninogenica, Prevotella nigrescens, Prevotella oralis, and Prevotella tannerae. Prevalence and risk analysis were performed using Student's t-test and Spearman's correlation. Among the Prevotella and Porphyromonas species detected in the periodontal lesions of sheep, P. melaninogenica (85.7%), P. buccae (64.3%), P. gingivalis (50%), and P. endodontalis (50%) were most prevalent. P. gingivalis (15%) and P. oralis (10%) prevailed in the gingival sulcus. P. gulae and P. tannerae were not detected in the 34 samples studied. Data evaluation by t-test verified that occurrence of P. asaccharolytica, P. endodontalis, P. gingivalis, P. buccae, P. intermedia, P. melalinogenica, and P. nigrescens correlated with sheep periodontitis. The findings of this study will be an important contribution to research on pathogenesis of sheep periodontitis and development of its control measures.

The microbiome of peri-implantitis sites and healthy dental sites was determined in subjects with a history of periodontitis. Ten systemically healthy non-smokers received periodontal treatment before implant placement. Plaque samples were collected from four sites of one implant with peri-implantitis (bone loss ≥2mm; PPD ≥6mm; BOP/suppuration; >1year loading) and from one dental site per quadrant with periodontal health (PPD ≤3mm, CAL <4mm, absence of BOP and bone loss). Following DNA extraction, the bacteria present in each sample were determined by high-throughput sequencing of the V3-V4 region of the 16S-rRNA gene using the Illumina-MiSeq platform. OTUs were picked using QIIME. Differences between dental and implant sites were determined using linear discriminant analysis, effect size and diversity analyses were conducted using PAST v3.02. The microbiomes of diseased samples were less diverse than those found in health, although disease was associated with a higher abundance of taxa relative to health. The genera Actinobacillus and Streptococcus were most closely associated with health, whereas Prevotella and Porphyromonas were most discriminative for disease. Other disease associated genera were Peptostreptococcus, Tannerella, Treponema, TG5 and Atopobium. Diseased peri-implant and healthy periodontal tissues in the same individual appear to harbour distinct microbiological ecosystems with putative periodontal pathogens predominating in the peri-implantitis-associated microbiome.

OBJECTIVE:To determine the composition of the microbiome of peri-implantitis sites and corresponding dental sites in subjects with a history of chronic periodontitis. DESIGN:Clinical and radiographic examination assessed the periodontal/peri-implant disease status. Plaque samples were collected from one diseased implant with peri-implantitis, functional for at least two years and healthy sites in ten non-smokers who had received periodontal treatment prior to implant placement. Following DNA extraction, the bacteria present in each sample were determined by high-throughput sequencing of V3-V4 region of the 16S rRNA gene using the Illumina MiSeq platform. OTUs were picked using QIIME. Differences between dental and implant sites were determined using linear discriminant analysis, effect size and diversity analyses were conducted using PAST v3.02. RESULTS:The microbiomes of healthy samples were more diverse than those found in disease, although disease was associated with a higher abundance of taxa relative to health. The genera Actinobacillus and Streptococcus were most closely associated with health, whereas Prevotella and Porphyromonas were most discriminative for disease. Synergistetes were highly associated with peri-implantitis. CONCLUSION:In patients with a history of periodontitis, putative periodontal pathogens prevailed in the microbiome of diseased implants. Diseased implants and corresponding healthy sites appear to have distinct microbiological ecosystems.

Feline odontoclastic resorptive lesion (FORL) and feline chronic gingivostomatitis (FCGS) are two of the most common diseases of the feline oral cavity. While evidence is emerging that FCGS is caused by gingival inflammation initiated and perpetuated by the oral microbiota, little is known in this regard for FORL. Feline calicivirus (FCV) has been associated with the presence of FCGS and is thought to play a role in the initiation of this disease. In this study, the incidence of FCV was investigated in cats with FORL and FCGS, and compared to unaffected controls. FCV was detected by viral culture. The incidence of FCV was as follows: 6 (24.0%) of 24 control cats, 9 (22.5%) of 40 cats with FORL and 15 (60.0%) of 25 cats with FCGS were positive for FCV. There was a significant difference in FCV incidence between all the groups (p=0.003) but none between the control group and the FORL group. However, significant differences were observed in the incidence of FCV between control and FCGS (p=0.010) and between FORL and FCGS (p=0.006). It is concluded that although FCV may be associated with FCGS, it appears unlikely to play a role in FORL.

PurposeMechanically ventilated patients are at risk for developing ventilator-associated pneumonia, and it has been reported that dental plaque provides a reservoir of respiratory pathogens that may aspirate to the lungs and endotracheal tube (ETT) biofilms. For the first time, metataxonomics was used to simultaneously characterize the microbiome of dental plaque, ETTs, and non-directed bronchial lavages (NBLs) in mechanically ventilated patients to determine similarities in respective microbial communities and therefore likely associations.Material and methodsBacterial 16S rRNA gene sequences from 34 samples of dental plaque, NBLs, and ETTs from 12 adult mechanically ventilated patients were analyzed.ResultsNo significant differences in the microbial communities of these samples were evident. Detected bacteria were primarily oral species (e.g., Fusobacterium nucleatum, Streptococcus salivarius, Prevotella melaninogenica) with respiratory pathogens (Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus pneumoniae, and Haemophilus influenzae) also in high abundance.ConclusionThe high similarity between the microbiomes of dental plaque, NBLs, and ETTs suggests that the oral cavity is indeed an important site involved in microbial aspiration to the lower airway and ETT. As such, maintenance of good oral hygiene is likely to be highly important in limiting aspiration of bacteria in this vulnerable patient group.