The Relationship between Uterine, Fecal, Bedding, and Airborne Dust Microbiota from Dairy Cows and Their Environment: A Pilot Study.

The aim of this study was to characterize uterine, fecal, bedding, and airborne dust microbiota from postpartum dairy cows and their environment. The cows were managed by the free-stall housing system, and samples for microbiota and serum metabolite assessment were collected during summer and winter when the cows were at one and two months postpartum. Uterine microbiota varied between seasons; the five most prevalent taxa were Enterobacteriaceae, Moraxellaceae, Ruminococcaceae, Staphylococcaceae, and Lactobacillaceae during summer, and Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, Moraxellaceae, and Clostridiaceae during winter. Although Actinomycetaceae and Mycoplasmataceae were detected at high abundance in several uterine samples, the relationship between the uterine microbiota and serum metabolite concentrations was unclear. The fecal microbiota was stable regardless of the season, whereas bedding and airborne dust microbiota varied between summer and winter. With regards to uterine, bedding, and airborne dust microbiota, Enterobacteriaceae, Moraxellaceae, Staphylococcaceae, and Lactobacillaceae were more abundant during summer, and Ruminococcaceae, Lachnospiraceae, Bacteroidaceae, and Clostridiaceae were more abundant during winter. Canonical analysis of principal coordinates confirmed the relationship between uterine and cowshed microbiota. These results indicated that the uterine microbiota may vary when the microbiota in cowshed environments changes.

Rumen fluid, feces, milk, water, feed, airborne dust, and bedding microbiota in dairy farms managed by automatic milking systems.

Microbiota of the gut, milk, and cowshed environment were examined at two dairy farms managed by automatic milking systems (AMS). Feed, rumen fluid, feces, milk, bedding, water, and airborne dust were collected and the microbiota on each was assessed by Illumina MiSeq sequencing. The most abundant taxa in feed, rumen fluid, feces, bedding, and water were Lactobacillaceae, Prevotellaceae, Ruminococcaceae, Ruminococcaceae, and Lactobacillaceae, respectively, at both farms. Aerococcaceae was the most abundant taxon in milk and airborne dust microbiota at farm 1, and Staphylococcaceae and Lactobacillaceae were the most abundant taxa in milk and airborne dust microbiota at farm 2. The three most prevalent taxa (Aerococcaceae, Staphylococcaceae, and Ruminococcaceae at farm 1 and Staphylococcaceae, Lactobacillaceae, and Ruminococcaceae at farm 2) were shared between milk and airborne dust microbiota. Indeed, SourceTracker indicated that milk microbiota was related with airborne dust microbiota. Meanwhile, hierarchical clustering and canonical analysis of principal coordinates demonstrated that the milk microbiota was associated with the bedding microbiota but clearly separated from feed, rumen fluid, feces, and water microbiota. Although our findings were derived from only two case studies, the importance of cowshed management for milk quality control and mastitis prevention was emphasized at farms managed by AMS.

Clinical risk factors for non-convulsive status epilepticus during emergent electroencephalogram.

PURPOSE: Emergent electroencephalograms (EmEEG) are performed to exclude non-convulsive status epilepticus (NCSE) but are resource-intensive. Prior studies have identified a seizure or seizures in the acute setting preceding the EmEEG request as a risk factor of NCSE but few other consistent clinical risk factors have been identified. We aimed to identify clinical risk factors for NCSE in EmEEGs METHODS: We conducted a retrospective analysis of consecutive patients who underwent EmEEG to exclude NCSE over a 20-month period. One blinded investigator extracted clinical information from patient case records using a standardized form. Patients were grouped using EmEEG results into those with and without NCSE. We analyzed differences between these two groups. RESULTS: A total of 2333 EEGs were performed over the study period, 215 (9.3%) were EmEEGs ordered to exclude NCSE. 21 patients (9.8%) of the 215 patients were found to have NCSE. Three independent clinical risk factors for NCSE were identified--seizure(s) in the acute setting, ocular movements (nystagmus and/or gaze deviation) and ongoing CNS infection. The presence of seizure(s) in the acute setting showed the highest adjusted odds ratio (OR=8.8, 95% CI 2.0-39.4, p=0.005). In addition, prevalence of NCSE increased as more clinical risk factors were present. CONCLUSION: Seizures in the acute setting, ocular movements and ongoing CNS infection are associated with NCSE. By using these risk factors at the bedside, clinicians can prioritize patients for EmEEG, recognizing that risk of NCSE increases as more clinical risk factors are present.

Identification of CYP3A4 and CYP2C8 as the major cytochrome P450 s responsible for morphine N-demethylation in human liver microsomes.

1. The aim was to identify the cytochrome P450 (CYP) enzymes responsible for the N-demethylation of morphine in vitro. 2. In human liver microsomes, normorphine formation followed Michaelis-Menten kinetics with mean Km and Vmax of 12.4 +/- 2.2 mM and 1546 +/- 121 pmol min(-1) mg(-1), respectively. In microsomes from a panel of 14 human livers phenotyped for 10 CYP enzymes, morphine N-demethylation correlated with testosterone 6beta-hydroxylation (r=0.91, p<0.001) and paclitaxel 6-alpha hydroxylation (r=0.72, p<0.001), two specific markers of CYP3A4 and CYP2C8, respectively. Normorphine formation decreased when incubated in the presence of troleandomycin or quercetin (by 46 and 33-36%, respectively), which further corroborates the contribution of CYP3A4 and CYP2C8. 3. Among eight recombinant human CYP enzymes tested, CYP3A4 and CYP2C8 exhibited the highest intrinsic clearance. More than 90% of morphine N-demethylation could be accounted for via the action of both CYP3A4 and CYP2C8. 4. The in vitro findings suggest that hepatic CYP3A4, and to a lesser extent CYP2C8, play an important role in the metabolism of morphine into normorphine.

Combination of a direct thrombin inhibitor and a platelet glycoprotein IIb/IIIa blocking peptide facilitates and maintains reperfusion of platelet-rich thrombus with alteplase.

We sought to determine the efficacy of the combination of argatroban, a direct thrombin inhibitor, and G4120, a platelet glycoprotein (GP) IIb/IIIa blocker, to enhance thrombolysis with alteplase. Platelet-rich thrombus in the rabbit arterial thrombosis model is relatively resistant to alteplase despite the addition of aspirin and heparin. The adjunctive use of either direct thrombin inhibitors or GP IIb/IIIa inhibitors in thrombolysis has been investigated with encouraging, but limited, success. The usefulness of combining both agents as adjunctive therapy to thrombolysis has not been fully explored. Following platelet-rich thrombus formation in the rabbit, argatroban (3 mg/kg), G4120 (0.5 mg/kg), G4120 plus heparin (200 U/kg), or G4120 plus argatroban were intravenously infused over 60 minutes. Alteplase was given as intravenous boluses (0.45 mg/kg) at 15-minute intervals up to 4 doses or until reperfusion. Blood flow and bleeding time were monitored for 2 hours. The combination of G4120 plus argatroban resulted in a persistent patency in 5 of 7 animals compared with 0 of 6 for argatroban alone (p=0.02), 1 of 6 for G4120 alone (p=0.08), and 2 of 6 for G4120 plus heparin (p=0.2). Although during the infusion the bleeding times were longer in the groups that received G4120 (26+/-7.7 minutes vs. 14+/-10 minutes, p<0.05), by the end of the experiment there were no statistically significant differences. Similarly, during the infusion the activated partial thromboplastin times (aPTT) was higher in groups that received heparin or argatroban (99+/-51 seconds vs. 32+/-7.6 seconds, p<0.001), but by the end of the experiment the aPTTs had returned to close to baseline in all groups except the G4120 plus heparin group. These results suggest that lysis of platelet-rich thrombus with alteplase requires the addition of both potent platelet and thrombin inhibitors. Specifically designed agents, G4120 and argatroban, are effective without additional increased risk for bleeding.

The effects of delta agonists on locomotor activity in habituated and non-habituated rats.

The effects of the delta agonists SNC80 and deltorphin II on ambulation and rearing activity were measured in habituated and non-habituated rats. SNC80 (30, 100, 200, 400 nmol, i.c.v.) and deltorphin II (3, 15, 30, 60 nmol, i.c.v.) induced similar, dose-dependent biphasic locomotor effects in non-habituated subjects. An initial decrease in exploratory activity was associated with anxiogenic signs such as pilo-erection, freezing behaviour and pupil dilation for each drug. Pre-treatment with the delta antagonist naltrindole (10 nmol, i.c.v.) inhibited the depressant effect, but not the subsequent stimulant effect, on locomotor activity in response to 30 nmol deltorphin II in this assay (P<0.05). In habituated rats, deltorphin II (0.03, 0.1, 0.3, 3 nmol, i.c.v.) caused significant, naltrindole-reversible increases in locomotor activity (P<0.05 for all doses) at 1,000-fold lower doses than those required for a similar response to SNC80 (10, 30, 100, 300 nmol, i.c.v.). Pharmacokinetic studies suggest that these compounds penetrate the brain to similar extents following i.c.v. injection. The substantial potency difference between deltorphin II and SNC80 in stimulating locomotor activity in habituated rats suggests pharmacological heterogeneity for these delta opioid receptor agonists.

High-performance liquid chromatographic assays with fluorometric detection for mivacurium isomers and their metabolites in human plasma.

Two high-performance liquid chromatographic assays coupled with fluorometric detection have been developed for the determination of mivacurium isomers (trans-trans, cis-trans and cis-cis) and their monoester and alcohol metabolites in human plasma. A novel solid-phase extraction procedure allowed good recovery of the mivacurium isomers (mean 98%) and their monoester metabolites (mean 83%), whereas the alcohol metabolites were analyzed after direct precipitation of plasma proteins. For all analytes, these assays proved to be sensitive (LOQ 3.9-15.6 ng/ml), reproducible (C.V. < 15%) and accurate (> 94%) over the therapeutic range of concentrations of mivacurium and its metabolites. These two methods were applied successfully to a pharmacokinetic study of mivacurium after a bolus dose of 0.15 mg/kg in anesthetized patients.

Peroxidase-catalyzed removal of phenols from coal-conversion waste waters.

A novel, enzymatic approach has been developed for the removal of phenols from coal-conversion aqueous effluents. Treatment with horseradish peroxidase and hydrogen peroxide precipitates 97 to 99 percent of the phenol in a wide range of pH and phenol concentrations; both model mixtures and real industrial waste-water samples have been treated successfully. Other pollutants, such as polychlorinated biphenyls, can be enzymatically coprecipitated with the phenols.