Genomic characterization of antifungal Acinetobacter bacteria isolated from the skin of the frogs Agalychnis callidryas and Craugastor fitzingeri.

Chytridiomycosis, a lethal fungal disease caused by Batrachochytrium dendrobatidis (Bd), is responsible for population declines and extinctions of amphibians worldwide. However, not all amphibian species are equally susceptible to the disease; some species persist in Bd enzootic regions with no population reductions. Recently, it has been shown that the amphibian skin microbiome plays a crucial role in the defense against Bd. Numerous bacterial isolates with the capacity to inhibit the growth of Batrachochytrium fungi have been isolated from the skin of amphibians. Here, we characterized eight Acinetobacter bacteria isolated from the frogs Agalychnis callidryas and Craugastor fitzingeri at the genomic level. A total of five isolates belonged to Acinetobacter pittii,Acinetobacter radioresistens, or Acinetobactermodestus, and three were not identified as any of the known species, suggesting they are members of new species. We showed that seven isolates inhibited the growth of Bd and that all eight isolates inhibited the growth of the phytopathogen fungus Botrytis cinerea. Finally, we identified the biosynthetic gene clusters that could be involved in the antifungal activity of these isolates. Our results suggest that the frog skin microbiome includes Acinetobacter isolates that are new to science and have broad antifungal functions, perhaps driven by distinct genetic mechanisms.

World Small Animal Veterinary Association Global Dental Guidelines.

Dental, oral, and maxillofacial diseases are some of the most common problems in small animal veterinary practice. These conditions create significant pain as well as localized and potentially systemic infection. As such, the World Small Animal Veterinary Association (WSAVA) believes that un- and under treated oral and dental diseases pose a significant animal welfare concern. Dentistry is an area of veterinary medicine which is still widely ignored and is subject to many myths and misconceptions. Effective teaching of veterinary dentistry in the veterinary school is the key to progression in this field of veterinary medicine, and to the improvement of welfare for all our patients globally. These guidelines were developed to provide veterinarians with the information required to understand best practices for dental therapy and create realistic minimum standards of care. Using the three-tiered continuing education system of WSAVA, the guidelines make global equipment and therapeutic recommendations and highlight the anaesthetic and welfare requirements for small animal patients. This document contains information on common oral and dental pathologies, diagnostic procedures (an easily implementable and repeatable scoring system for dental health, dental radiography and radiology) and treatments (periodontal therapy, extractions). Further, there are sections on anaesthesia and pain management for dental procedures, home dental care, nutritional information, and recommendations on the role of the universities in improving veterinary dentistry. A discussion of the deleterious effects of anaesthesia free dentistry (AFD) is included, as this procedure is ineffective at best and damaging at worst. Throughout the document the negative effects of undiagnosed and/or treated dental disease on the health and well-being of our patients, and how this equates to an animal welfare issue, is discussed.

Oral and Dental Examination Findings in European Polecats (Mustela putorius).

Dental and oral diseases in the domestic ferret have been reported, but comparison with their closest wild relative, the European polecat (Mustela putorius), is lacking. Dental and orofacial pathology was evaluated by means of visual examination and dental radiographical analysis of 234 museum skulls. Most of these (70.5%) originated from Austria and the remainder came from seven other European countries. Linguoverted mandibular second incisor teeth were noted in 77% of the skulls. There were 7,268 (91.4% of all potential) teeth available for examination. Few (0.3%) were presumed congenitally missing and 0.5% were presumed missing by acquired means. Supernumerary teeth were present in 2.6% of the skulls. Most (65.6%) of the maxillary first molar teeth were three rooted with the palatal root protruding into the orbit. Most (67.2%) of the mandibular first molar teeth were three rooted with the smaller third root being positioned in the furcation of the two larger roots. Thirty-one percent of the teeth were four rooted, with the smaller fourth root also being positioned in the furcation of the two largest roots. Attrition/abrasion was shown by 17.2% of the teeth in 47% of the skulls. Periodontitis was recognized in 57.6% of the teeth in 91.9% of the skulls and dental fractures were noted in 7.2% of the teeth in 62% of the skulls. Radiographical evidence of endodontic disease was found in 25% of the fractured teeth. Tooth resorption, in the form of external inflammatory resorption associated with endodontic disease, was found in 0.1% of teeth in 2.1% of skulls. Extrusion affected 44% of the canine teeth. Lesions consistent with infection by the helminth Troglotrema acutum were identified in 33.3% of the skulls. Oral and dental disease was significant and varied, often consistent with reports of the species domestic counterpart - the ferret. Comparison with other mustelids, notably the beech marten (Martes foina), showed both clear consistencies and discrepancies, dependent on the type of pathology.

Oral and Dental Examination Findings in Beech Martens (Martes foina).

Detailed clinical and radiographical descriptions of oral/dental pathology in the beech marten (Martes foina) are lacking. In the present study, skulls of M. foina from a museum collection (n = 109) were examined macroscopically according to predefined criteria and full-mouth dental radiographs were obtained. Occlusion was diagnosed as 'scissor bite' in 22.5% of evaluated skulls, 77% of evaluated skulls showed 'level bite' of incisors and 9.2% of evaluated skulls had 'open bites'. In three skulls, class III malocclusion was diagnosed. Linguoversion of the mandibular second incisor teeth was noted in all skulls. Of the maximum possible number of teeth (i.e. full set of teeth in each skull), 90.4% were available for examination, 6.4% were missing artifactually, 2.0% were absent presumably congenitally and in 1.2 % the absence was presumably acquired. In three skulls supernumerary teeth were noted, all being incisor teeth. In four skulls, unerupted maxillary canine teeth were noted and in one skull odontodysplasia of the maxillary canine teeth was diagnosed. All other teeth were considered normal in morphology. The number of roots per tooth varied in the mandibular second premolar, maxillary first molar and mandibular second molar teeth. The most common dental pathology was attrition/abrasion with 72 skulls (66%) and 857 teeth (24.1% of evaluated teeth) affected, followed by periodontal disease affecting 52 (47.7%) of skulls and 773 teeth (18.9% of evaluated teeth). Dental fractures were present in 49 (45%) skulls and 148 teeth (3.8% of evaluated teeth). Radiographically evident periapical lesions were detected in 11 (10.1%) skulls and 18 teeth (0.5% of evaluated teeth). Other rare abnormal findings included tooth resorption, enamel hypoplasia/hypocalcification, fenestrations at palatal root of maxillary first molar tooth and different bony changes.

Oral health and systemic inflammatory, cardiac and nitroxid biomarkers in hemodialysis patients.

BACKGROUND: Periodontal diseases have systemic inflammatory effects and have been adversely associated with cardiovascular diseases, which are also the most frequent cause of death in the end-stage renal disease. The aim of this cross-sectional study was to investigate the oral health and serum biomarkers among the hemodialysis (HD) patients in Slovenia. MATERIAL AND METHODS: 111 HD patients were periodontally examined and their sera were assayed for C reactive protein (CRP), cardiac troponin T (TnT), nitrite/nitrate (NOx) and antibody levels to A. actinomycetemcomitans and P. gingivalis. The association of oral health with systemic response was analyzed with Kruskal-Wallis test, Fisher's exact test and multivariate linear regression. RESULTS: Bleeding on probing without periodontal pockets was present in 5.2%, calculus without periodontal pockets in 42.1%, shallow periodontal pockets in 39.5% and deep periodontal pockets in 13.2% of dentate patients. There were 28.8% edentulous participants. 63.1% of the patients had CRP levels higher than 3 mg/L and 34.2% higher than 10 mg/L. TnT was detectable in all participants, with 25.2% exhibiting levels higher than 100 ng/L. The median level of NOx was 43.1 µmol/L. Participants with higher CRP were more likely to be edentulous and have higher TnT levels. A direct association of oral health with TnT or NOx was not detected. CONCLUSIONS: HD patients in Slovenia have compromised oral health and increased serum inflammatory and cardiac biomarkers. Edentulousness was an independent predictor for the increased CRP, indicating a need for improved dental care to retain the teeth as long as possible.

Oral and dental diseases in a population of domestic ferrets (Mustela putorius furo).

OBJECTIVE: Domestic ferrets have been used for many purposes, but recently their popularity as companion animals has increased greatly. However, data on their oral and dental diseases are rare. The objective of this study was therefore to describe oral and dental diseases in a population of client-owned domestic ferrets. METHODS: In this cross-sectional clinical study, detailed oral and dental examination and full-mouth dental radiographs were performed in 57 client-owned ferrets. RESULTS: Variations in occlusion and number of roots per tooth were noted in comparison with previously published literature on ferrets. Periodontal disease, attrition/abrasion and dental fractures, especially of the canine teeth, were commonly observed. Periapical disease associated with dental fractures, malocclusion, tooth resorption and neoplasia was uncommon. CONCLUSION AND CLINICAL RELEVANCE: This study indicates that thorough oral and dental examination supported by dental radiography under general anaesthesia should be performed in domestic ferrets as a part of regular veterinary care.

Discrimination of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex species by Fourier transform infrared spectroscopy.

The main goal of this work was to assess the ability of Fourier transform infrared spectroscopy with attenuated total reflectance (FTIR-ATR) to discriminate between the species of the Acinetobacter calcoaceticus-Acinetobacter baumannii (Acb) complex, i.e. A. baumannii, A. nosocomialis, A. pittii, A. calcoaceticus, genomic species "Between 1 and 3" and genomic species "Close to 13TU". A total of 122 clinical isolates of the Acb complex previously identified by rpoB sequencing were studied. FTIR-ATR spectra was analysed by partial least squares discriminant analysis (PLSDA) and the model scores were presented in a dendrogram form. This spectroscopic technique proved to be effective in the discrimination of the Acb complex species, with sensitivities from 90 to 100%. Moreover, a flowchart aiming to help with species identification was developed and tested with 100% correct predictions for A. baumannii, A. nosocomialis and A. pittii test isolates. This rapid, low cost and environmentally friendly technique proved to be a reliable alternative for the identification of these closely related Acinetobacter species that share many clinical and epidemiological characteristics and are often difficult to distinguish. Its validation towards application on a routine basis could revolutionise high-throughput bacterial identification.

Oral papillary squamous cell carcinoma in twelve dogs.

Papillary squamous cell carcinoma (PSCC) is a distinct histological subtype of oral squamous cell carcinoma (SCC), described in both dogs and man. In dogs, PSCC has long been considered a malignant oral tumour of very young animals, but it has recently been reported to occur in adult dogs as well. The aim of this study was to describe the major clinicopathological characteristics of canine oral PSCC (COPSCC). Twelve dogs diagnosed with COPSCC were included in this retrospective study (1990-2012). The majority (75%) of the dogs were >6 years of age (median age 9 years). All tumours were derived from the gingiva of dentate jaws, with 66.7% affecting the rostral aspects of the jaws. The gross appearance of the lesions varied, with one having an intraosseous component only. The majority (91.7%) of the tumours were advanced lesions (T2 and T3), but no local or distant metastases were noted. Microscopically, two patterns were seen: (1) invasion of bone forming a cup-shaped indentation in the bone or a deeply cavitating cyst within the bone (cavitating pattern), (2) histologically malignant growth, but lack of apparent bone invasion (non-cavitating pattern). The microscopical appearance corresponded to imaging findings in a majority of cases, with cavitating forms presenting with a cyst-like pattern of bone loss or an expansile mass on imaging and non-cavitating forms showing an infiltrative pattern of bone destruction on imaging. These features suggest two distinct biological behaviours of COPSCC.

Periodontal disease, periodontal treatment and systemic nitric oxide in dogs.

Thirty-two client-owned dogs treated for periodontal disease were divided in group 1 if no periodontitis, group 2 if ≤25%, and group 3 if >25% of the teeth present were affected with periodontitis. Blood was tested before and 2 weeks after periodontal therapy for nitrosyl hemoglobin (HbNO), plasma nitrite/nitrate (NOx) and 3-nitrotyrosine (NT) levels. No HbNO was detected in any of the animals tested. There was no significant difference in the NOx plasma levels within each group or across the groups before and after the treatment, but a noticeable increase in NOx plasma levels was observed in group 3 after the treatment. Plasma NT was detected in only one third of the animals. NO levels varied greatly across individual dogs. The data are suggestive of an overall increase in systemic NO response 2 weeks after periodontal treatment in dogs with advanced periodontal disease, but the response is greatly individually-dependent.

Erythema multiforme and epitheliotropic T-cell lymphoma in the oral cavity of dogs: 1989 to 2009.

OBJECTIVES: To describe the clinical signs, major laboratory findings, diagnostic procedures and outcome in dogs with erythema multiforme or epitheliotropic T-cell lymphoma of the oral cavity. METHODS: Retrospective analysis identified 14 client-owned dogs with erythema multiforme or epitheliotropic T-cell lymphoma of the oral cavity. Histological changes were combined with immunohistochemistry and clonality testing data in selected cases, and a clinical follow-up was acquired. RESULTS: Ulcerative stomatitis with no significant or only minor abnormalities on haematology and serum biochemistry panels was common. Histological features were diagnostic in seven cases. The use of immunohistochemistry distinguished the two diseases in an additional three cases. In four cases, the diagnosis remained questionable, and clonality testing for T-cell receptor gamma gene rearrangement helped confirm erythema multiforme in one case. Clinical follow-up revealed erythema multiforme in two and epitheliotropic T-cell lymphoma in one of the remaining questionable cases. CLINICAL SIGNIFICANCE: Erythema multiforme and epitheliotropic T-cell lymphoma may affect, but are rarely limited to the oral cavity of dogs, and they usually present as stomatitis. Histological features alone are not always diagnostic. Immunohistochemistry and clonality testing may assist in the differentiation between the two, but in ambiguous cases, repeated biopsy and clinical follow-up are essential.

Variability in the concentrations of volatile metabolites emitted by genotypically different strains of Pseudomonas aeruginosa.

AIMS: To characterize the volatile metabolites produced by genotypically diverse strains of Pseudomonas aeruginosa in order to evaluate their potential for use as biomarkers of lung infection in noninvasive breath analysis. METHODS AND RESULTS: Volatile organic compounds (VOCs) emitted from 36 clinical strains of Ps. aeruginosa (belonging to different multilocus sequence types) cultured in liquid and on solid media were analysed by gas chromatography mass spectrometry (GC-MS) and selected ion flow tube mass spectrometry (SIFT-MS). Several previously identified VOCs were detected, including ethanol, acetone, 2-butanone, 2-pentanone, isoprene, aminoacetophenone, dimethyl sulphide, dimethyl disulphide, dimethyl trisulphide and methyl thiocyanate. Additionally, significant production of 3-methyl-butanone, acetophenone, methylthioacetate and methyl thiobutanoate was observed for the first time in this study. SIFT-MS quantifications of VOCs showed high variability between genotypically distinct strains. CONCLUSIONS: The data obtained indicate that the production rates of the volatile biomarkers of Ps. aeruginosa vary by two orders of magnitude between different strains cultured under the same conditions. Similar variability was observed for both liquid and solid media. SIGNIFICANCE AND IMPACT OF THE STUDY: Inter-strain genotypic variability strongly influences the concentrations of the volatile biomarkers from Ps. aeruginosa. A group of several biomarkers quantified in real time in exhaled breath may thus provide a more valuable indicator of the course of pulmonary infections compared to a single biomarker.

Histological subtypes of oral non-tonsillar squamous cell carcinoma in dogs.

Several histological subtypes and grades of oral squamous cell carcinoma (SCC) are described in human literature and these subtypes have distinct morphological features and biological behaviour. This retrospective study (1990-2010) included 84 dogs diagnosed with SCC of the oral cavity and oropharynx, excluding the tonsils. Sixty-nine of the SCCs (82.1%) were further diagnosed as conventional SCC (CSCC) (33 [47.8%] well-differentiated, 31 [44.9%] moderately-differentiated and five [7.3%] poorly-differentiated), five (5.95%) each as papillary SCC and basaloid SCC, three (3.6%) as adenosquamous carcinoma and two (2.4%) as spindle cell carcinoma. Compared with the general hospital population, neutered female dogs, dogs aged 10 to <15 years, English springer spaniels and Shetland sheepdogs were overrepresented. The majority (78.1%) of SCCs were proliferative with or without associated ulceration, although no significant association was observed between the gross appearance and different SCC subtypes. 71.4% of SCCs were located in dentate jaws; however, well-differentiated CSCC more often affected the tongue and other non-dentate mucosal surfaces (P=0.0022). No significant association was found between any of the SCC subtypes and tumour-associated inflammation (TAI), perineural and lymphovascular invasion (PNI, LVI), or between gross appearance of the tumour and tumour location, PNI, LVI or TAI or PNI, LVI, TAI and tumour location.

Expression of cytokeratins in the epithelium of canine odontogenic tumours.

Odontogenic tumours are considered to be relatively rare; however, several histologically distinct types have been identified in dogs. The more common canine odontogenic tumours are peripheral odontogenic fibroma and canine acanthomatous ameloblastoma. The expression of cytokeratins (CKs) has been established for the human dental germ and odontogenic tumours. The aim of the present study was to describe the immunohistochemical expression of a panel of CKs in the epithelium of the canine dental germ, normal gingiva and odontogenic tumours arising in this species. Samples from 20 odontogenic tumours, 12 tooth germs and three normal gingival tissues were obtained. Each sample was stained with haematoxylin and eosin and subjected to immunohistochemistry for CK expression. The typical expression pattern of CKs in the odontogenic epithelium and gingiva of dogs was CK14 and CK5/6. CKs 7, 8, 18 and 20 were generally absent from the canine dental germ, gingiva and odontogenic tumours. Dogs and man therefore exhibit similar CK expression in the odontogenic epithelium.

Analysis of immune cells within the healthy oral mucosa of specific pathogen-free cats.

The oral mucosa is an important interface for host-environment interactions. Based on previous studies, it is generally accepted that the cellular compartments of the oral immune system comprise organized mucosal-associated lymphoid tissues as well as diffusely and focally distributed T- and to lesser extent B-lymphocytes, oral mucosal Langerhans cells (OMLC), macrophages and mast cells. However, a comprehensive quantification of the cellular elements in the oral mucous membranes of the cat has not been reported. The aim of this study was to provide a comprehensive analysis of the immune cell compartments in the oral mucous membranes and anatomically related tissues of healthy cats. Multiple biopsies of the oral mucous membranes and related tissues were obtained from four specific pathogen-free cats for histological and immunohistochemical assessment of lymphocyte subsets, OMLC, macrophages and mast cells. T-lymphocyte subsets, OMLC, mast cells and macrophages were present in varying frequencies among the tissue compartments of the feline oral cavity. B-lymphocytes were not identified in any of the examined tissues except the tonsils and mandibular lymph nodes. Lymphocytic aggregates (follicles) were found in the palatoglossal folds and the gingiva. We describe the topographical distribution of various leucocyte subsets in the normal healthy feline oral mucosa, and demonstrate regional differences in the distribution of these cells.

Chronic ingestion of Porphyromonas gingivalis induces systemic nitric oxide response in mice.

INTRODUCTION: Porphyromonas gingivalis induces nitric oxide (NO) production in various cells, systemic NO elevation being expected in chronic oral challenge. METHODS: Groups of BALB/c mice were inoculated orally with either live P. gingivalis ATCC 33277 or sterile broth on days 0, 2 and 4, with or without later administration of the inducible nitric oxide synthase (iNOS) inhibitor 1400W. Plasma and tissues were harvested on day 42 for assays of tumor necrosis factor-alpha (TNF-alpha), nitrite and nitrate (NOx) and tissue NO, or histology and iNOS immunohistochemistry. RESULTS: No signs of gingivitis were observed, but plasma NOx was significantly elevated (P = 0.028) as was TNF-alpha (P = 0.079) in P. gingivalis-inoculated animals compared with controls, NOx being reduced when 1400W was used. NO production in organs showed a similar trend, with significant elevation in liver (P = 0.017) and kidneys (P = 0.027), whereas concomitant treatment of inoculated animals with 1400W caused significant reductions in NO in aorta (P = 0.008) and kidneys (P = 0.046). Sham-inoculated 1400W-treated animals had significantly increased plasma NOx (P = 0.004) and liver NO (P = 0.04). NOx in plasma correlated significantly with NO production in lungs (0.35, P = 0.032) and kidneys (0.47, P = 0.003). Immunohistochemistry demonstrated iNOS activity in many tissues in all groups. CONCLUSION: Repeated oral administration of P. gingivalis induced systemic NO and NOx production in mice, probably by activating iNOS as suggested by the response to 1400W.

Single gavage with Porphyromonas gingivalis reduces acute systemic nitric oxide response in mice.

INTRODUCTION: Porphyromonas gingivalis, an important periodontal pathogen, can also induce host responses in distant tissues. P. gingivalis induces nitric oxide (NO) production in immune system cells and non-immune system cells, therefore NO might be involved in an acute systemic host response. METHODS: Eighteen female BALB/c mice were perorally inoculated with 10(8) colony-forming units live P. gingivalis ATCC 33277. Plasma nitrite and nitrate (NOx) and NO production in lungs, aorta, heart, liver, spleen, kidneys, and brain were measured at intervals after inoculation and compared with levels in 11 control animals. RESULTS: NOx levels were significantly (P = 0.017) lower at 7, 13, and 25 h after P. gingivalis inoculation. A similar trend in NO production occurred in most tested organs, but never reached statistical significance. The correlation between NOx in plasma and NO in liver was positive (Spearman correlation coefficient = 0.81, P = 0.0025) and marginal for kidney (0.58, P = 0.059). CONCLUSION: Single peroral inoculation of mice with P. gingivalis reduces the acute systemic NO response. As NO is important for host defense, the reduction of NO levels after exposure is likely to delay the host response, increasing the chances that infection with P. gingivalis will become established.

Total intravenous anaesthesia with propofol or propofol/ketamine in spontaneously breathing dogs premedicated with medetomidine.

The cardiorespiratory parameters, the depth of anaesthesia and the quality of recovery were evaluated in six spontaneously breathing dogs that had been premedicated with medetomidine (40 microg/kg, supplemented with 20 microg/kg an hour later), administered with either propofol (1 mg/kg followed by 0.15 mg/kg/minute, intravenously), or with ketamine (1 mg/kg followed by 2 mg/kg/hour, intravenously) and propofol (0.5 mg/kg followed by 0.075 mg/kg/minute, intravenously). The dogs' heart rate and mean arterial blood pressure were higher and their minute volume of respiration and temperature were lower when they were anaesthetised with propofol plus ketamine, and a progressive hypercapnia leading to respiratory acidosis was more pronounced. When the dogs were anaesthetised with propofol/ketamine they recovered more quickly, but suffered some unwanted side effects. When the dogs were anaesthetised with propofol alone they recovered more slowly but uneventfully.

Prevalence of Acinetobacter baumannii and other Acinetobacter spp. in faecal samples from non-hospitalised individuals.

In total, 226 individuals from the community were investigated for faecal carriage of Acinetobacter spp. by broth enrichment culture, followed by growth on blood agar and/or Leeds Acinetobacter Medium (LAM). Acinetobacter baumannii was isolated on both LAM and blood agar from one of 100 specimens in the UK and one of 126 specimens in The Netherlands. The predominant species were Acinetobactor johnsonii and genomic sp. 11, which were cultured from 22 and five specimens, respectively. A. baumannii did not seem to be widespread in the faecal flora of individuals in the community.