Dextranase enzyme and Enterococcus faecium probiotic have anti-biofilm effects by reducing the count of bacteria in dental plaque in the oral cavity of dogs.

OBJECTIVE: Periodontal disease is a common clinical complication and has a negative impact on the quality of life and the welfare of companion dogs. Periodontal disease occurs when pathogenic bacteria are accumulated in the gingival sulcus, which favors biofilm formation. The oral health of dogs can be significantly compromised by dental plaque accumulation. Thus, this investigation demonstrates the effect of Enterococcus faecium probiotic, dextranase enzyme, and their combination on dental biofilm in the oral cavity of dogs. ANIMALS: The 30 dogs were referred to Polyclinic with no oral ulcers, severe periodontitis, and internal diseases. PROCEDURES: Dextranase enzyme, E faecium probiotic, and their combination were administered in the oral cavity of dogs. Microbiological samples were obtained from tooth surfaces and gums before and after intervention with the substances. Bacterial colonies were enumerated by using a colony counter. Also, Porphyromonas gingivalis hmuY gene expression was evaluated by reverse transcription quantitative real-time PCR analysis. RESULTS: The total colony count of the bacterial culture indicated that the dextranase enzyme, E faecium probiotic, and their combination significantly reduced the total bacteria count in the oral cavity. Moreover, in the reverse transcription quantitative real-time PCR analysis it was observed that using the combination of E faecium probiotic and dextranase enzyme decreases the hmuY gene expression of P gingivalis bacteria. CLINICAL RELEVANCE: The results clearly indicated that the dextranase enzyme and E faecium probiotic could be used as preventive agents to reduce oral biofilm in dogs. Furthermore, no side effects were observed while using these substances.

In vitro evaluation of the antibacterial effect of Brazilian red propolis ethanol extract in the prevention of periodontal disease in dogs.

Dental plaque (DP) is found on the surface of teeth and comprises a community of microorganisms that form a structured biofilm. Bacteria present in DP are potential periodontal pathogens when there is an imbalance in the healthy oral environment, and are precursors of periodontal disease (PD). In dogs, the treatments, such as mechanical removal, are difficult and expensive to apply. Therefore, in order to seek new therapeutic alternatives to control dental plaque in dogs, Brazilian red propolis ethanol extract (RPEE) was tested to evaluate its antibacterial effect on bacteria isolated from DP of dogs without PD. DP was collected from the supragingival dental surfaces of 10 dogs. Bacterial isolates of DP were identified by PCR and sequencing of 16S rDNA gene. The RPEE was obtained using the ultrasound ethanol extraction technique, and the chemical composition was obtained by HPLC-DAD and UV-spectrophotometry. In total, 29 different bacteria belonging to five genera were identified. Formononetin, biochanin A, liquiritigenin and daidzein were the major constituents of the RPEE. The cytotoxic effect showed cell viability after 24 h above 50 % at all concentrations evaluated. The minimum inhibitory concentration was between 37.5 and 150.0 µg/mL for all bacterial isolates. The minimal bactericidal concentration was between 150 and 1200 µg/mL for Gram-positive and 300-1200 µg/mL for Gram-negative bacteria. The results are promising and suggest that RPEE has significant antibacterial potential against the bacteria present in the DP of healthy dogs. Although further studies are still needed, the results suggest RPEE might be safely used in the prevention of periodontal disease.

Effect of TrisEDTA and Chlorhexidine 0.12% on an In Vitro-Defined Biofilm Representing the Subgingival Plaque Biofilm of the Dog.

This study was designed to investigate the effects of chlorhexidine 0.12%, TrisEDTA (tromethamine ethylenediamintetraacetic acid), and a combination of chlorhexidine 0.12% and TrisEDTA on an in vitro plaque biofilm model comprised of three bacterial species commonly found in canine subgingival plaque. Porphyromonas gulae, Actinomyces canis, and Neisseria canis were grown in a biofilm on polished hydroxyapatite coated titanium alloy pucks for 72 h prior to exposure to one of four test solutions: TrisEDTA, chlorhexidine 0.12%, a combination of TrisEDTA and chlorhexidine 0.12%, or sterile deionized water as a control. Following exposure to the test solution, a sample was collected of the biofilm either immediately or following 24 h of additional incubation in a broth medium. Lower numbers of CFU/mL of Porphyromonas gulae resulted when the biofilm was treated with a solution of chlorhexidine 0.12% and TrisEDTA compared to with chlorhexidine 0.12% alone, TrisEDTA alone, or the control and so this solution can be said to be synergistic against Porphyromonas gulae in this controlled in vitro model. Greater reductions in the numbers of CFU/mL of Actinomyces canis and Neisseria canis resulted from treatment with chlorhexidine 0.12% alone than if treated with the combination of TrisEDTA and chlorhexidine 0.12%. When treated biofilm samples were allowed 24 h of additional growth in fresh media, greater variance resulted and this variance highlights the complex dynamics involved in bacterial growth within a biofilm.

Comparison of a Vegetable-Based Dental Chew to 2 Other Chews for Oral Health Prevention.

Giving dental chews to dogs is part of the passive homecare that helps prevent the formation of plaque and tartar. The objectives of these studies were to assess the effectiveness of a vegetable-based dental chew (VF) to maintain oral health, and to compare it to 2 different reference chews (RC) with a proven effectiveness. The first study was conducted on 45 small dogs (<10 kg) and the second on 60 larger dogs (15-30 kg) who were randomly assigned to 3 different groups. During 30 days, one group received no chew (control) while the second and third group received either one RC (RC1 or RC2) or one VF per day. All dogs had their teeth scaled on Day 0. On Day 30, scores were given for plaque and calculus. Gingival parameters were also assessed. Statistical analysis (analysis of variance and Tukey tests ± Bonferroni's adjustment) were performed to compare groups with α set at .05 for significance.The 3 types of chews were found to be efficacious to reduce plaque and calculus formation and the gingival bleeding compared to control (P < .05). There was no significant difference between RCs and VF in both trials except for the gingival bleeding parameters which showed a greater improvement with VF. Therefore, daily administration of the VF is effective to reduce plaque and calculus formation and gingival bleeding and has a better efficacy on gingival bleeding than the other reference products tested. It can therefore be used with confidence at home for preventative dental care.

Porphyromonas spp., Fusobacterium spp., and Bacteroides spp. dominate microbiota in the course of macropod progressive periodontal disease.

Macropod progressive periodontal disease (MPPD) is a necrotizing, polymicrobial, inflammatory disease commonly diagnosed in captive macropods. MPPD is characterized by gingivitis associated with dental plaque formation, which progresses to periodontitis and then to osteomyelitis of the mandible or maxilla. However, the underlying microbial causes of this disease remain poorly understood. In this study, we collected 27 oral plaque samples and associated clinical records from 22 captive Macropodidae and Potoroidae individuals that were undergoing clinical examination at Adelaide and Monarto Zoos in South Australia (15 healthy, 7 gingivitis and 5 periodontitis-osteomyelitis samples). The V3-V4 region of the 16S ribosomal RNA gene was sequenced using an Illumina Miseq to explore links between MPPD and oral bacteria in these animals. Compositional differences were detected between the microbiota of periodontitis-osteomyelitis cases compared to healthy samples (p-value with Bonferroni correction < 0.01), as well as gingivitis cases compared to healthy samples (p-value with Bonferroni correction < 0.05) using Permutational Multivariate Analysis of Variance (PERMANOVA). An overabundance of Porphyromonas, Fusobacterium, and Bacteroides taxa was also identified in animals with MPPD compared to healthy individuals using linear discriminant analysis effect size (LEfSe; p = < 0.05). An increased abundance of Desulfomicrobium also was detected in MPPD samples (LEfSe; p < 0.05), which could potentially reflect differences in disease progression. This is the first microbiota analysis of MPPD in captive macropods, and these results support a polymicrobial pathogenesis of MPPD, suggesting that the microbial interactions underpinning MPPD may be more complex than previously documented.

Ozone Therapy in the Prevention of Dental Plaque Formation in Dogs.

This study aims to assess ozonized mineral oil ointment application as an antiplaque therapy for dogs. Domestic healthy dogs received dental scaling and polishing under general anesthesia. Under standard feeding and homecare during 7 days, 20 dogs were randomly placed into 2 different groups for dental treatment. The control group (CG) was given a single placebo application and the ozone group (O3G) received daily ozonized ointment application. The average age (CG = 4.4; O3G = 5.7 years old), body weight (CG = 15.7; O3G = 15.3 kg) and the gingivitis index obtained on the first day (D0) allowed initial homogeneity between the groups. The dental plaque index, including clinical and computerized analysis on the seventh day, was obtained from the buccal aspect of specific dental locations. Both analyses revealed significant statistical association between daily application of ozone and antiplaque effect. There was no evidence of toxicity during the study. These results suggest that ozone therapy may be an efficient adjuvant to conventional periodontal treatment in decreasing initial dental plaque formation.

Comparison of subgingival and gingival margin plaque microbiota from dogs with healthy gingiva and early periodontal disease.

The purpose of the investigation was to determine whether canine gingival margin (GM) plaque is a reliable surrogate for subgingival (SG) plaque from a microbial community (microbiota) perspective. SG and GM plaque samples were collected from 381 dogs visiting pet hospitals in the USA, China and Thailand. Dogs with clinically healthy gingivae through to early periodontitis were included in the study. The samples were subject to next generation Illumina sequence analysis to allow microbiota comparisons to be made between the two plaque sources. Overall, the SG and GM samples indicated commonality via the majority community that were shared between them; health associations led to the identification of some significant taxa-specific differences. GM microbiota exhibited lower variability and diversity and were shown to reflect a sub-population of those associated with SG plaque. Both plaque niches, however, demonstrated similar changes in microbial signatures with health and early periodontal disease and did not indicate divergent trends. The key, most abundant microbiota of GM plaque strongly reflect those observed with SG plaque across health and early periodontitis. Microbiota in plaque from above the gum line may therefore be employed as a biomarker of oral health. This opens up the potential to use plaque, sampled from conscious dogs, to define oral health status and improve the diagnosis, treatments and interventions for periodontal disease.

Orally administered ovine serum immunoglobulins modulate dental plaque in cats.

The effects of orally administered ovine serum immunoglobulin on dental plaque and associated oral immunity in cats were investigated. The two treatment groups consisted of 1) cats that were fed unsupplemented kibble (control diet) and 2) cats that were fed the same kibble but coated with a freeze-dried ovine serum immunoglobulin preparation (ovine Ig) (test diet). The adult cats were randomly allocated to one of the two diets (n = 15) and received their respective kibble for a 28-day experimental period. When compared to the ovine Ig-supplemented kibble, cats consuming the unsupplemented kibble had significantly (p < 0.05) higher dental plaque scores. Cat IgA and IgG concentrations in the saliva and serum were significantly (p < 0.05) higher for cats fed the unsupplemented kibble when compared to cats receiving the ovine Ig supplement. Similarly, myeloperoxidase activity in the saliva was significantly (p < 0.05) higher for cats fed the unsupplemented kibble when compared to cats receiving the Ig-supplement. Orally administered ovine serum Ig positively influenced oral health and oral immunity in cats as evidenced by preventing an increase of dental plaque formation, salivary and serum IgA and IgG concentrations and salivary myeloperoxidase activity.

Evaluation of Efficacy of a Dental Chew to Reduce Gingivitis, Dental Plaque, Calculus, and Halitosis in Toy Breed Dogs.

The aim of this study was to determine the clinical efficacy of a dental chew with mechanical and chemical properties in 2 toy dog breeds. Eight Yorkshire terriers and 9 Chihuahuas participated in a crossover design trial. Gingivitis, dental plaque, calculus, and volatile sulfur compounds (VSC) in the breath were assessed after 4 weeks and 9 weeks, respectively. When dogs were fed 1 dental chew per day they had significantly reduced gingivitis (-20%, P < .001), accumulation of plaque (-15%, P < .001), calculus (-35%, P = .001), and VSC concentration (-19%, P < .001) compared to when receiving no chew. A significant breed effect was observed on all the average dental indices and VSC concentration.

Effect of feeding a daily oral care chew on the composition of plaque microbiota in dogs.

The objective of this study was to investigate the influence of daily feeding of an oral care chew on the composition of canine supragingival plaque microbiota. Twelve beagle dogs were recruited to the randomized cross-over study. The dogs were fed one of two dietary regimes, both consisting of a commercially available wet and dry diet mix, either with or without daily supplementation with an oral care chew. After each 28-day test phase, supragingival plaque samples were collected and processed via Illumina sequencing to determine the microbiota composition. A comparative analysis of bacterial species associated with health and periodontal disease, identified from prior clinical studies, revealed differences between the dietary regimes. Consumption of the daily oral care chew, resulted in a significant increase in proportion of 6 health associated taxa but only 3 disease associated taxa compared to no chew. In contrast, 8 disease and 1 health associated taxa showed increased proportions for no chew versus the oral care chew. Daily feeding of the oral care chew tested in this study has therefore been shown to increase the proportion of health associated bacteria, over bacteria associated with periodontal disease, in supragingival plaque compared to no chew. By influencing plaque microbiota towards a bias for health associated bacteria, feeding of the oral care chew provides a means to reduce the prevalence of bacterial species shown to be associated with periodontal disease in dogs.

A microbiome analysis of equine peripheral dental caries using next generation sequencing.

BACKGROUND: Although, peripheral caries (PC) affects almost half of UK horses, no comprehensive microbiological study has been performed on this disorder. As a high proportion of oral bacteria cannot be conventionally cultured, molecular microbiological techniques such as Next Generation Sequencing are required to examine the complex oral bacteria community. OBJECTIVES: To identify the microbiota involved in equine PC, including comparing microbiota at the more commonly and severely affected three caudal cheek teeth with the less commonly affected three rostral cheek teeth. STUDY DESIGN AND METHODS: Equine dental plaque samples were collected from the palatal aspects of cheek teeth of 63 horses. DNA was isolated and amplified using PCR, targeting the V4 region of the 16S rRNA gene and Next Generation Sequencing of these gene amplicons was performed. The acquired data were processed and analysed using Mothur and R. RESULTS: Streptococcus species was the genus most commonly associated with equine PC, whereas Gemella species was the genus most associated with the control group. In a further analysis where the rostral and caudal cheek teeth were compared with each other and with the control group. Veillonella species was the most commonly associated genus with PC of the rostral cheek teeth, Streptococcus species was the most associated genus with the caudal cheek teeth, and Corynebacterium with the control group. MAIN LIMITATIONS: Some bacteria can have multiple heterogeneous copies of the 16S rRNA gene, which can affect the estimation of their relative abundance. CONCLUSIONS: Similar to caries studies in other species, acidogenic and aciduric microorganisms including Streptococcus species were found to be associated with equine peripheral caries.

Chemical and structural composition of black pigmented supragingival biofilm of bovines with periodontitis / Composição química e estrutural do biofilme supragengival pigmentado de bovinos com periodontite

Bovine periodontitis is a multifactorial disease primarily associated with a potentially pathogenic microbiota housed in the oral biofilm of animals. Biofilms are organized structures, in which the constituents coexist in symbiosis, already described as a predisposing factor to periodontitis in other species. The objective of the present study was to characterize the structure and chemical aspects of the bovine black pigmented supragingival biofilm using scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS), respectively, and determine its relationship with bovine periodontitis. Eleven premolar teeth from different animals were evaluated; five non-pigmented samples and six samples with black pigmented biofilms were initially submitted to SEM, and three areas of these samples were selected for EDS. The structure of the pigmented biofilm was more complex and irregular because of a higher content of mineral elements. The semi-quantitative EDS data indicated an association of iron (p<0.014) and magnesium (p<0.001) with the occurrence of periodontitis, whereas carbon, phosphorus, calcium, manganese, sodium, and potassium were not associated with the disease. Carbon (p<0.039), manganese (p<0.007), and iron (p<0.015) were associated with pigmentation, whereas phosphorus, calcium, and magnesium were not associated with it. Spearman correlation test showed the relationships between calcium and phosphorus, and iron and silicon. The strong association of iron in the pigmented supragingival biofilm and with the occurrence of periodontitis suggests the presence of microorganisms that use this element in their metabolism and that are also associated with bovine periodontitis. This study suggests that the pigmented deposits in the crown of the teeth of cattle are an true biofilm with the deposition of iron, and it indicates that the presence of iron and magnesium in these formations may be involved in the metabolism of some microorganisms associated with the etiology of bovine periodontitis.(AU)

A periodontite bovina é uma infecção multifatorial associada primariamente à microbiota potencialmente patogênica presente no biofilme bucal. Biofilmes são estruturas organizadas, nas quais os constituintes convivem em simbiose, descritos em outras espécies como um fator predisponente à periodontite. O objetivo do presente estudo foi caracterizar estrutural e quimicamente o biofilme supragengival pigmentado de preto em bovinos, utilizando-se as técnicas de microscopia eletrônica de varredura (MEV) e espectroscopia de dispersão de energia (EDS), respectivamente, correlacionando os elementos identificados à ocorrência de periodontite e pigmentação. Foram avaliados 11 dentes primeiro-molares; cinco amostras sem pigmentação visível e seis amostras com biofilme pigmentado de preto, que foram submetidas inicialmente à MEV; posteriormente foram selecionadas três áreas aleatórias de cada dente para realização da EDS. A estrutura do biofilme pigmentado revelou formações irregulares e mais complexas, provavelmente devido ao maior acúmulo de elementos minerais. Os resultados semi-quantitativos da EDS apontaram associações entre a presença de ferro (p<0,014) e magnésio (p<0,001) com a ocorrência de periodontite. Carbono, fósforo, cálcio, manganês, sódio e potássio não apresentaram associação com a periodontite. Em relação à pigmentação, carbono (p<0,039), manganês (p<0,007) e ferro (p<0,015) foram os elementos estatisticamente significantes, enquanto fósforo, cálcio e magnésio não apresentaram associação com a pigmentação. O teste de correlação de Spearman demonstrou associações entre os elementos cálcio e fósforo, e ferro e silício. A forte associação do ferro presente no biofilme supragengival com a ocorrência de periodontite, sugere a presença de micro-organismos que utilizam este elemento em seu metabolismo e que possivelmente tenham envolvimento com o desenvolvimento da periodontite bovina. Os resultados inéditos do presente trabalho sugerem que os depósitos pigmentados que se formam na coroa dos dentes de bovinos são um biofilme verdadeiro com deposição de ferro, e indicam que a presença de ferro e magnésio nestas formações pode estar envolvida no metabolismo de alguns dos principais micro-organismos associados à etiologia da periodontite bovina.(AU)

Two Randomized Trials Demonstrate Lactic Acid Supplementation in Pet Food Inhibits Dental Plaque, Calculus, and Tooth Stain in Cats.

BACKGROUND: Dental concerns are some of the most common health problems affecting companion animals. A variety of foods, treats, and chews comprising different mechanical and chemical technologies have been investigated as a means of promoting oral health. Here, we investigate the chemical technology, lactic acid added to a commercially available food, for its ability to inhibit dental plaque, calculus, and tooth stain accumulation in cats. METHODS: Two separate feeding trials assessed the utility of a nutritionally complete feline maintenance food supplemented with lactic acid to reduce oral substrate accumulation (dental plaque, calculus, and tooth stain) in cats. After a calibration study identified high and low dental plaque formers, 45 cats were randomized to 1 of 2 test groups (food with 1.2% lactic acid supplementation) or control (food without lactic acid supplementation) groups, stratified based on their calibration scores. Data were collected on a monthly basis for 3 months. The second study randomly assigned 24 cats to either the test or control groups for 1 year, with data collected at the 6- and 12-month time points. RESULTS: In the 3-month study, reductions in dental plaque, calculus, and tooth stain accumulations were observed at the 2-month assessment in both test groups compared with control (P < .05 for test group 2). The 1-year study showed that these reductions in oral substrate accumulation persisted through the 6- and 12-month time points (P < .05). CONCLUSIONS: Taken together, these studies demonstrate that lactic acid supplemented at 1.2% in a feline maintenance food significantly inhibits oral substrate accumulation.

Development of dental hygiene gum for cats considering their anatomical features of dentition.

The purpose of this study was to develop effective dental hygiene chews for cats based on the anatomical features of the dentition and patterns of chewing motion. All cats were volunteered for dental prophylaxis followed by dental impressions using yellow stone and alginate under general anesthesia. Twenty parameters related to dentition were defined in order to compare dental impressions using digital caliper. For the chewing motion study, patterns of chewing motion were identified based on recordings made with a digital camera. Ten cats (4 domestic shorthairs, 2 Russian blues, 1 American shorthair, 1 Persian, 1 Turkish Angora, and 1 Devon Rex) were recruited for the study. The parameters related to teeth and oral size were similar among the studied cats. Chewing motion can be described as more of a guillotine-like motion rather than a crushing motion, with cats chewing 3-7 times before swallowing. The chewing pattern of cats involves shearing for a short period of time followed by immediate swallowing. Therefore, the overall size of the dental hygiene chew could be determined based on the measurement of the oral size for inducing chewing. The surface details of the dental hygiene chew could be designed for prevention and removal of dental calculus and plaque in cats considering the anatomical teeth parameters. Dental hygiene chews customized for cats considering the different anatomical features of their teeth might be effective for oral care.

Prospective randomised blinded clinical trial assessing effectiveness of three dental plaque control methods in dogs.

OBJECTIVES: To assess the effectiveness of three commonly used methods of controlling dental plaque accumulation within a general practice setting. MATERIALS AND METHODS: A blinded study, block randomising dogs at recruitment to receive one of three treatments to start after a routine scale and polish procedure: once daily tooth brushing using a toothbrush and veterinary toothpaste, once daily administration of a single dental hygiene chew or a prescription dental diet. Blinded scoring of plaque accumulation, both coverage and thickness, was performed at the end of the 6-week study. RESULTS: A total of 22 dogs completed the study, which simultaneously assessed the effectiveness of the three means of plaque control. Daily tooth brushing was shown to be more than three times as effective at controlling plaque accumulation compared to using a daily dental chew or dental diet. The dental plaque scores for the diet and dental chews were not significantly different from one another and there was much greater variability in the mouth scores for these two groups of dogs compared to the tooth brushing group. CLINICAL SIGNIFICANCE: The study design has been shown to be an effective means of assessing plaque accumulation over a 6-week period using a clean mouth model. Secondly, this study adds to the evidence which shows that daily tooth brushing is the most effective single method of reducing plaque accumulation and optimising dogs' oral health.

Potential of two delivery systems for nisin topical application to dental plaque biofilms in dogs.

BACKGROUND: Periodontal disease (PD) is caused by the development of a microbial biofilm (dental plaque) in the periodontium, affecting approximately 80% of dogs. Several bacterial species present in the canine oral cavity can be implicated in the development of this disease, including Enterococcus spp. To decrease antibiotic administration, a possible control strategy for dog's enterococcal PD may involve the use of the antimicrobial peptide (AMP) nisin. Nisin's inhibitory activity was evaluated against a collection of previously characterized enterococci obtained from the oral cavity of dogs with PD (n = 20), as well as the potential of a guar-gum gel and a veterinary toothpaste as topical delivery systems for this AMP. The Minimum Inhibitory (MIC) and Bactericidal Concentrations (MBC) and the Minimum Biofilm Eradication (MBEC) and Inhibitory Concentrations (MBIC) were determined for nisin and for the supplemented guar-gum gel. For the supplemented veterinary toothpaste an agar-well diffusion assay was used to evaluate its inhibitory potential. RESULTS: Nisin was effective against all isolates. Independently of being or not incorporated in the guar-gum gel, its inhibitory activity on biofilms was higher, with MBIC (12.46 ± 5.16 and 13.60 ± 4.31 µg/mL, respectively) and MBEC values (21.87 ± 11.33 and 42.34 ± 16.61 µg/mL) being lower than MIC (24.61 ± 4.64 and 14.90 ± 4.10 µg/mL) and MBC (63.09 ± 13.22 and 66.63 ± 19.55 µg/mL) values. The supplemented toothpaste was also effective, showing inhibitory activity against 95% of the isolates. CONCLUSIONS: The inhibitory ability of nisin when incorporated in the two delivery systems was maintained or increased, demonstrating the potential of these supplemented vehicles to be applied to PD control in dogs.

The occurrence of Treponema spp. in gingival plaque from dogs with varying degree of periodontal disease.

Periodontal disease is common in dogs and is initiated by gingival plaque composed of several hundred bacterial species. Some of these species have specifically been pointed out as potential periodontal pathogens, such as Treponema spp. Treponema spp. are difficult to culture and therefore the majority have been detected by culture-independent methods, such as PCR (Polymerase Chain Reaction). This leaves many Treponema spp. uncharacterized and unnamed. In this study, treponemes were investigated in gingival plaque from dogs with varying degree of periodontal disease with the aim to describe their occurrence and diversity in dogs. The methods used were culture, phase-contrast microscopy, PCR targeting the 16SrRNA-tRNAIle intergenic spacer region (ISR2), sequencing of the ISR2 and phylogenetic analysis. Treponema spp. were detected in samples from 10 out of 11 dogs and isolates were obtained from six dogs. Both healthy and periodontitis affected dogs were Treponema positive. Phylogenetic analysis, based on ISR2 sequences, revealed a large diversity of treponemes in the study population that were found to be distributed mainly in two groups, corresponding to the human oral treponeme phylogroups II (Treponema denticola) and IV (Treponema maltophilum) genetic groups. They were generally more distantly related to other treponemes in these groups. Treponemes from dogs with periodontitis and dogs with mild gingivitis without periodontitis did not differ in any obvious way. The results indicate that several phylotypes of oral treponemes are common in dogs regardless of periodontal status.

The effect of dietary corn oil and fish oil supplementation in dogs with naturally occurring gingivitis.

The aim of this randomized, double-blinded, placebo-controlled study was to evaluate if downregulation of the inflammatory response due to ingestion of high levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) can slow down gingivitis development, and thus delay the progression of periodontal disease (PD) in dogs. To this aim, 44 client-owned adult dogs (>1 and <8 years old) with naturally occurring PD (stages 1 and 2) were submitted to a plaque, gingivitis and calculus scoring followed by a dental cleaning procedure and collection of blood samples. The animals were then fed a canine adult maintenance diet, supplemented with either corn oil (0.00 g EPA and 0.00 g DHA) or fish oil (1.53 g EPA and 0.86 g DHA, both per 1,000 kcal ME) over the following 5 months. At the end of this period, the PD scoring and the blood sampling were repeated. The animals consuming fish oil had higher plasma levels of the longer chain (C ≥ 20) omega 3 fatty acids (p < 0.01) and similar plasma levels of alpha-linolenic acid (p = 0.53), omega 6 fatty acids (p > 0.63) and C reactive protein (p = 0.28) then the ones consuming corn oil. There were no differences between fish oil and corn oil diet supplementation on plaque (18.2 vs. 17.8, p = 0.78), calculus (10.1 vs. 11.5, p = 0.18) or gingivitis (19.3 vs. 19.0, p = 0.77) indexes. The authors conclude that supplementation with EPA + DHA does not slow down progression of PD in dogs.

Assessment of dental plaque coverage by Quantitative Light-induced Fluorescence (QLF) in domestic short-haired cats.

Dietary means of reducing plaque and calculus deposits are frequently sought for the maintenance of oral health in cats and dogs. In the development of such products sensitive, reliable, reproducible methods of measuring plaque and calculus are key. The aim of this study was to assess Quantitative Light-induced Fluorescence (QLF™) for the detection of dental plaque coverage in cats compared to the modified Logan and Boyce technique. The techniques were utilised in a crossover study, which compared two diets for their effect on plaque deposition in a cohort of 24 adult cats. Analysis of the effect of diet on plaque coverage by both the modified Logan and Boyce technique and QLF showed a significant effect of feeding regime (p=0.024 and p≤0.0001, respectively) with good agreement between the techniques in the percentage reduction of plaque accumulation. A within study assessment of QLF demonstrated excellent intra-operator repeatability (coefficient of variation 2.2%). Similarly, inter-operator reproducibility was also good (coefficient of variation 2.3%). A retrospective analysis, using the data to estimate the sample size required for at least 90% power to detect a 15% difference between treatments in a two-way crossover study, established that 10 cats would be sufficient for plaque measurement by QLF, while assessment by the modified Logan and Boyce method required over 30 cats. QLF was determined to be a reliable, reproducible method for the assessment of plaque deposition in cats and requires fewer subjects for the detection of differences between treatment effects compared to the modified Logan and Boyce method.

The Prevalence of Canine Oral Protozoa and Their Association with Periodontal Disease.

Periodontal disease is one of the most important health concerns for companion animals. Research into canine forms of periodontitis has focused on the identification and characterization of the bacterial communities present. However, other microorganisms are known to inhabit the oral cavity and could also influence the disease process. A novel, broad spectrum 18S PCR was developed and used, in conjunction with next-generation sequencing analyses to target the identification of protists. Trichomonas sp. and Entamoeba sp. were identified from 92 samples of canine plaque. The overall prevalence of trichomonads was 56.52% (52/92) and entamoebae was 4.34% (4/92). Next-generation sequencing of pooled healthy, gingivitis, early-stage periodontitis, and severe periodontitis samples revealed the proportion of trichomonad sequences to be 3.51% (health), 2.84% (gingivitis), 6.07% (early periodontitis), and 35.04% (severe periodontitis), respectively, and entamoebae to be 0.01% (health), 0.01% (gingivitis), 0.80% (early-stage periodontitis), and 7.91% (severe periodontitis) respectively. Both genera of protists were statistically associated with plaque from dogs with periodontal disease. These findings provide the first conclusive evidence for the presence of oral protozoa in dog plaque and suggest a possible role for protozoa in the periodontal disease process.