Microbiology insights into boosting salivary defences through the use of enzymes and proteins.

OBJECTIVES: To assess the effects of enzymes and proteins in toothpastes to boost salivary defences and reduce oral bacteria growth and viability. METHODS: An in vitro study to measure levels of hypothiocyanite in saliva after treatment with a toothpaste containing enzymes and proteins (colorimetric assay). A randomised, crossover in vivo study measuring, with biochemical assays, the effect of a toothpaste containing enzymes and proteins on the levels of hydrogen peroxide and lysozyme in saliva. In vitro studies to measure, using fluorescent dyes, the effects of enzymes and proteins on bacterial membrane integrity. In vitro microbiology studies measuring the effects of enzymes and proteins on planktonic bacterial growth, in vitro studies in single and multispecies biofilms measuring the effect of toothpaste with enzymes and proteins compared to control toothpaste. RESULTS: Levels of hypothiocyanite, hydrogen peroxide and lysozyme are boosted after application of a toothpaste containing enzymes and proteins. The enzymes and proteins adversely affect the bacterial membrane integrity in Streptococcus mutans and growth of planktonic S. mutans and Fusobacterium nucleatum. In a single species biofilm model the viability of S. mutans was significantly reduced and in a 7 species biofilm model bacterial viability was reduced for a biofilm grown on a pellicle pre-treated with toothpaste containing enzymes and proteins. CONCLUSIONS: The use of a toothpaste containing enzymes and proteins can boost the natural salivary defences by increasing the levels of lysozyme and hydrogen peroxide in vivo and hypothiocyanite in vitro and reduce the growth and viability of oral bacteria in microbiological models. CLINICAL SIGNIFICANCE: The subtle effects reported here of enzymes and proteins in toothpastes on oral bacteria are consistent with both the gum health benefits reported for such toothpastes containing enzymes and proteins by Daly [1] and Pedersen [2] and the rebalancing of the oral microbiome reported by Adams [3].

Association between characteristics of pain and stiffness and the functional status of patients with incident polymyalgia rheumatica from primary care.

This paper aims to examine the relationship between different characteristics of pain and stiffness and the functional status of patients with newly diagnosed polymyalgia rheumatica (PMR). Baseline analysis of an inception cohort study was conducted. Patients aged ≥18 years, with a new diagnosis of PMR were recruited from 382 English general practices. Participants were mailed a baseline questionnaire, including separate pain and stiffness manikins and numerical rating scales (NRS), a question on their ability to raise their arms above their head and the modified Health Assessment Questionnaire (mHAQ) to examine participants' functional status. Linear regression analysis, reported as regression co-efficients (95% confidence intervals (95% CI)), was used to assess the association of pain and stiffness with function, initially unadjusted and then adjusted for age, gender, deprivation status, smoking status, BMI, anxiety and depression. Six hundred fifty two patients responded to the baseline survey (88.5%). The majority (88.2%) reported no, or mild impairment in their functional status. Adjusted linear regression analysis demonstrated that high (NRS ≥8) pain (0.20 (95% CI 0.10-0.28)) or stiffness (0.18 (0.09-0.26)) ratings, an increasing number of sites of pain (0.18 (0.06-0.29)) or stiffness (0.19 (0.08-0.31)) and shoulder pain (0.18 (0.05-0.31)), stiffness (0.10 (0.01-0.20)) and difficulty raising arms above one's head (0.19 (0.10-0.28)) were all associated with increased functional impairment. The majority of newly diagnosed PMR patients reported no or minimal functional difficulty. However, those who experience severe or widespread pain or stiffness often have significant functional limitation in performing their daily activities and may be a subset worthy of additional focus in primary care.

Aptamer based peptide enrichment for quantitative analysis of gonadotropin-releasing hormone by LC-MS/MS.

Over recent years threats to racing have expanded to include naturally occurring biological molecules, such as peptides and proteins, and their synthetic analogues. Traditionally, antibodies have been used to enable detection of these compounds as they allow purification and concentration of the analyte of interest. The rapid expansion of peptide-based therapeutics necessitates a similarly rapid development of suitable antibodies or other means of enrichment. Potential alternative enrichment strategies include the use of aptamers, which offer the significant advantage of chemical synthesis once the nucleic acid sequence is known. A method was developed for the enrichment, detection and quantitation of gonadotropin-releasing hormone (GnRH) in equine urine using aptamer-based enrichment and LC-MS/MS. The method achieved comparable limits of detection (1 pg/mL) and quantification (2.5 pg/mL) to previously published antibody-based enrichment methods. The intra- and inter-assay precision achieved was less than 10% at both 5 and 20 pg/mL, and displayed a working dynamic range of 2.5-100 pg/mL. Significant matrix enhancement (170 ± 8%) and low analytical recovery (29 ± 15%) was observed, although the use of an isotopically heavy labelled GnRH peptide, GnRH (Pro(13)C5,(15)N), as the internal standard provides compensation for these parameters. Within the current limits of detection GnRH was detectable up to 1h post administration in urine and identification of a urinary catabolite extended this detection window to 4h. Based on the results of this preliminary investigation we propose the use of aptamers as a viable alternative to antibodies in the enrichment of peptide targets from equine urine.

Searching for new markers of endogenous steroid administration in athletes: "looking outside the metabolic box".

A simple means of detecting the abuse of steroids that also occur naturally is a problem facing doping control laboratories. Specific markers are required to allow the detection of the administration of these steroids. These markers are commonly measured using a set of data obtained from the screening of samples by gas chromatography-mass spectrometry (GC-MS). Doping control laboratories further need to confirm identified abuse using techniques such as gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). An interesting urinary species was found while following the pharmacokinetics and changes to the steroid profile from single and multiple oral doses of the International Olympic Committee/World Anti Doping Agency (IOC/WADA) prohibited substance, dehydroepiandrosterone (DHEA). The urine samples collected from the administration studies were subject to GC-MS and GC-C-IRMS steroid analysis following cleanup by solid phase extraction techniques. A useful urinary product of DHEA administration was detected in the urine samples from each of the administration studies and was identified by GC-MS experiments to be 3alpha,5-cyclo-5alpha-androstan-6beta-ol-17-one (3alpha,5-cyclo). This compound occurs naturally but the concentrations of 3alpha,5-cyclo were elevated following both the single DHEA administration (up to 385 ng/mL) and multiple DHEA administrations (up to 1240 ng/mL), in relation to those observed prior to these administrations (70 and 80 ng/mL, respectively). A reference distribution of urine samples collected from elite athletes (n = 632) enabled the natural concentration range of 3alpha,5-cyclo to be established (0-280 ng/mL), with a mean concentration of 22 ng/mL. Based on this an upper 3alpha,5-cyclo concentration limit of 140 ng/mL is proposed as a GC-MS screening marker of DHEA abuse in athletes. GC-C-IRMS analysis revealed significant 13C depletion of 3alpha,5-cyclo following DHEA administration. In the single administration study, the delta13C value of 3alpha,5-cyclo changed from -24.3 per thousand to a minimum value of -31.1 per thousand at 9 h post-administration, before returning to its original value after 48 h. The multiple administration study had a minimum delta13C 3alpha,5-cyclo of -33.9 per thousand during the administration phase in contrast to the initial value of -24.2 per thousand. Preliminary studies have shown 3alpha,5-cyclo to most likely be produced from DHEA sulfate found at high levels in urine. The complementary use of GC-MS and GC-C-IRMS to identify new markers of steroid abuse and the application of screening criteria incorporating such markers could also be adapted by doping control laboratories to detect metabolites of androstenedione, testosterone and dihydrotestosterone abuse.

Nutrient processing capacity of a constructed wetland in western Ireland.

In Ireland, constructed wetland systems are increasingly being used to perform tertiary treatment on municipal waste effluent from small towns and villages located in areas whose receiving waters are deemed sensitive. The bedrock formation in the west of Ireland is primarily karst limestone and where the overburden-soil cover is very shallow, such waters are highly sensitive to pollution sources, as little or no natural attenuation and/or treatment will occur. Constructed wetland technology has been seen to offer a relatively low-cost alternative to the more conventional tertiary treatment technologies, particularly when dealing with low population numbers in small rural communities. This paper examines the waste treatment performance, in terms of nutrient (P and N) reduction, of a recently constructed surface-flow wetland system at Williamstown, County Galway, Ireland. Performance evaluation is based on more than two years of water quality and hydrological monitoring data. The N and P mass balances for the wetland indicate that the average percentage reduction over the two-year study period is 51% for total N and 13% for total P. The primary treatment process in the wetland system for suspended solids (between 84 and 90% reduction), biological oxygen demand (BOD) (on average, 49% reduction), N, and P is the physical settlement of the particulates. However, the formation of algal bloom during the growing season reduces the efficiency of the total P removal.

Blood supply to the periosteum of the canine tibia.

A study was undertaken to determine which arteries supply the periosteum of the normal adult canine tibia. The vessels were found to arise from locations distinct from the muscular vascularization. The venous drainage of the tibial periosteum had a satellite relationship with the arteries. On the surface of the periosteum, the vessels formed rectangular patterns. All surfaces of the distal end of the tibia seemed to have less distinct anastomoses of the transverse periosteal twigs, compared with the proximal end of the tibia.

Progressive oropharyngeal dysfunction in a dog.

A dog with an acquired, progressive oropharyngeal dysphagia also had a myopathy-neuropathy. It was clinically similar to criocopharyngeal achalasia, and could easily have been confused with it, even with fluoroscopic evaluation. Conservative medical therapy was instituted since cricopharyngeal myotomy could have caused severe aspiration and death. Resolution was apparently due to anti-inflammatory therapy.

Radiographic examination of mandibular lesions in barren-ground caribou.

Dental anomalies were observed in 43 of 1,226 barren-ground caribou (Rangifer tarandus groenlandicus) taken between 1966 and 1968. In five of these 43 animals, the mandibles had deformities which radiography showed to be the result of dental abscesses in four cases and probably of a trauma in the other. The absence of actinomycotic lesions of the jaw bones of these 1,226 animals, and of more than 500 examined previously, indicates that "lumpy jaw" is rare in barren-ground caribou. The authors suggest the use of radiography to determine the nature of bone growth on skeletal remains, in the absence of soft tissues for examination for Actinomyces, either microscopically or by cultural methods.

Changes in the acid base status of sheep anaesthetised with a combination of atropine sulphate acepromazine and ketamine hydrochloride.

pH, PaCO2, PaO2, standard bicarbonate, base excess and reduced pH were measured in sheep before and at regular intervals after administration of ketamine with and without atropine and acepromazine premedication. A decrease in pH and PaO2 and a rise in PaCO2 was observed 15 minutes after administration of ketamine. Administration of atropine with and without acepromazine had no significant effect on pH, PaCO2 and PaO2. The values for standard bicarbonate, base excess and reduced pH were not significantly affected. This indicates that minor changes observed in pH, PaCO2 after ketamine administration are compensated for by the healthy animal's blood buffer system.

Polydactylism in a white-tailed deer from eastern Ontario.

A polydactylous right front foot was collected from a white -tailed deer (Odocoileus virginianus) fawn in eastern Ontario, Canada. We determined by gross visual and radiographic examinations that the polydactylous foot had a complete extra digit in the position of the ancestral 1st digit. This is the first documented account of a perfect polydactylous condition in a North American cervid.