Dysbiosis and predicted function of dental and ruminal microbiome associated with bovine periodontitis.

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.

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)

Efficacy of virginiamycin for the control of periodontal disease in calves / Eficácia da virginiamicina no controle de doença periodontal em bezerros

Periodontal diseases are multifactorial infectious processes caused by complexes of microorganisms, with damage to health, production, and animal welfare. The aim of the present study was to evaluate the efficacy of virginiamycin in the prevention and control of two early forms of periodontal disease: gingivitis and necrotizing gingivitis. Ten weaned calves, aged four to six months, were permanently kept in a single lot under the same rotational grazing regime in a newly reformed area of Panicum maximum. Five of the calves were orally administered 340mg of virginiamycin (Virginiamycin Group) daily for a period of 18 weeks, while the remaining five calves (Control Group) remained under the same food management but did not receive virginiamycin. During this period, animals underwent 18 weekly evaluations regarding periodontal health, with monitoring and recording of clinical parameters of the eight deciduous incisor teeth on the labial and lingual faces. At approximately two-week intervals, nine collections of subgingival sulcus material from five sites of the four right incisor teeth of each animal were performed and subjected to microbiological evaluation using polymerase chain reaction with primers of 25 microorganisms considered potentially pathogenic. After 1440 periodontal clinical evaluations of incisor teeth of the 10 calves, a total of 395 episodes of gingivitis were recorded, of which 267 occurred in the Control Group and 128 in the Virginiamycin Group. Similarly, 89 episodes of necrotizing gingivitis were recorded; 58 in the Control Group and 31 in the Virginiamycin Group. Comparison of between-group means found significant differences for teeth with gingivitis and necrotizing gingivitis (t test; p<0.05). The total number of teeth with gingivitis (p<0.01) and necrotizing gingivitis (p<0.01) in Control Group was significantly higher than that of gingivitis (p<0.01) and necrotizing gingivitis (p<0.05) in the Virginiamycin Group. There was a positive correlation between total occurrence of gingivitis and necrotizing gingivitis in the Virginiamycin Group by Pearson's test. Virginiamycin had a protective effect on treated animals compared with the Control Group (OR = 0.36: CI (95%) = 0.27-0.43). In the Control Group, Actinomyces israelli (4.74%), domain Archaea (1.58%), Eikenella corrodens (1.05%), Fusobacterium nucleatum (27.37%), class Mollicutes (5.26%); Porphyromonas endodontalis(5.26%); Porphyromonas gulae(0.53%), Prevotella buccae (6.32%), Prevotella loescheii (3.68%), Prevotella nigrescens (8.42%), Prevotella oralis (1.58%), Tannerella forsythia (0.53%), and Treponema denticola (4.21%) were detected at healthy sites, and gingivitis or necrotizing gingivitis samples. In the Virginiamycin Group, A. israelli (3.41%), domain Archaea (0.98%), F. nucleatum (9.27%), class Mollicutes(4.39%), P. endodontalis (4.39%), P. gulae (0.49%), P. buccae (8.29%), P. loescheii (6.83%), P. nigrescens (15.61%), P. oralis (1.46%), Selenomonas sputigena (0.49%), T. forsythia (0.49%), and T. denticola (2.44%) were detected. In conclusion, virginiamycin administered at a dosage of 340mg/animal/day significantly reduced the occurrence of gingivitis and necrotizing gingivitis in cattle maintained on reformed pastures, and was revealed to have action against periodontal bacterial microbiota considered to be potentially pathogenic.(AU)

As doenças periodontais são processos infecciosos multifatoriais causados por complexos de micro-organismos, que provocam danos à saúde, produção e ao bem-estar animal. O objetivo do presente estudo foi o de avaliar a eficácia da virginiamicina na prevenção e controle de duas formas de doença periodontal; a gengivite e a gengivite necrosante. Assim, dez bezerros desmamados, com idade entre 4 e 6 meses, foram mantidos permanentemente em lote único e sob o mesmo regime de pastejo rotacionado em área reformada de Panicum maximum. Cinco bezerros receberam via oral 340mg de virginiamicina (Grupo Virginiamicina) diariamente, por um período de dezoito semanas, enquanto o Grupo Controle permaneceu sob o mesmo manejo alimentar, mas sem receber a virginiamicina. No período, os animais passaram por 18 avaliações semanais quanto à saúde periodontal, com monitoramento e registro dos parâmetros clínicos dos oito dentes incisivos decíduos, nas suas faces labial e lingual. Em intervalos aproximadamente quinzenais foram realizadas nove coletas de material do sulco subgengival de cinco sítios de quatro dentes incisivos direitos de cada animal para avaliação microbiológica, com o emprego da reação em cadeia da polimerase e com iniciadores de 25 micro-organismos considerados potencialmente patogênicos. Ao final das 1440 avaliações clínicas periodontais dos dentes incisivos dos dez bezerros, pôde-se registrar um total de 395 episódios de dentes com gengivite, nos quais 267 foram registrados no Grupo Controle e 128 no Grupo Virginiamicina. De forma semelhante, do total de 89 registros de gengivite necrosante, 58 foram no Grupo Controle e 31 no Grupo Virginiamicina. Na comparação entre médias dos grupos as diferenças encontradas para dentes com gengivite e gengivite necrosante foram significativas pelo teste t (p<0,05). Assim, o total de dentes com gengivite (p<0,01) e gengivite necrosante (p<0,01) no Grupo Controle, foi significativamente superior ao de gengivite (p<0,01) e gengivite necrosante (p<0,05) do Grupo Virginiamicina. Houve correlação positiva entre o total de ocorrência de gengivite e gengivite necrosante no Grupo Virginiamicina pelo teste de Pearson. A virginiamicina possuiu um efeito protetor nos animais tratados em comparação com o controle (OR = 0,36: IC (95%) = 0,27-0,43). Na avaliação microbiológica do Grupo Controle foram detectados nas amostras de sítios sadios, com gengivite ou com gengivite necrosante Actinomyces israelli (4,74%), domínio Archaea (1,58%), Eikenella corrodens (1,05%), Fusobacterium nucleatum (27,37%), classe Mollicutes (5,26%), Porphyromonas endodontalis (5,26%), Porphyromonas gulae (0,53%), Prevotella buccae (6,32%), Prevotella loescheii (3,68%), Prevotella nigrescens (8,42%), Prevotella oralis (1,58%), Tannerella forsythia (0,53%) e Treponema denticola (4,21%). Enquanto no Grupo Virginiamicina foram detectados: A. israelli (3,41%), domínio Archaea (0,98%), F. nucleatum (9,27%), classe Mollicutes (4,39%), P. endodontalis (4,39%), P. gulae (0,49%), P. buccae (8,29%), P. loescheii (6,83%), P. nigrescens (15,61%), P. oralis (1,46%), Selenomonas sputigena (0,49%), T. forsythia (0,49%) e T. denticola (2,44%). Em conclusão, a virginiamicina administrada na dosagem de 340mg/animal/dia reduziu significativamente a ocorrência da gengivite e gengivite necrosante em bovinos mantidos em pastos reformados e revelou ter ação frente à microbiota bacteriana periodontal considerada potencialmente patogênica.(AU)

Black-pigmented anaerobic bacteria associated with ovine 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.