Introduction of new multidrug-resistant Salmonella enterica strains into commercial dairy herds.

A longitudinal observational study of 59 dairy herds was conducted in Washington State to estimate the rate of introduction of new multidrug-resistant (MDR) Salmonella enterica strains onto commercial dairy herds. Samples were collected on these herds over 7 visits separated by intervals of 2 to 4 mo over a period of 15 to 21 mo. Samples were cultured for Salmonella spp. and serogroup, serovar, and antimicrobial susceptibility patterns were identified for MDR Salmonella isolates. Fingerprinting generated by pulsed-field gel electrophoresis (PFGE) using XbaI restriction enzyme digestion generated genotyping profiles for all MDR isolates identified in the study. The rate of new MDR Salmonella strain introduction was 0.9 per herd-year (95% confidence interval: 0.6-1.4). The rates for the most commonly introduced MDR Salmonella serovars were 0.4/herd-year for Typhimurium, 1.2/herd-year for Newport, and 0.1/herd-year for Dublin. Thirty-three of 59 herds (56%) had at least one new MDR Salmonella introduction during the study period. The number of new MDR Salmonella strains acquired by dairy herds ranged from zero to 8. Thirteen of the 59 herds had a history of clinical salmonellosis. Among these 13 herds, 6 herds acquired new MDR Salmonella strains, although these strains were different than historical clinical strains. These data indicate that acquisition of new MDR Salmonella strains by dairy herds was a common event in participating herds, although the number of strains introduced varied greatly among herds.

Metabolism of [3H]pentosan polysulfate sodium (PPS) in healthy human volunteers.

Pentosan polysulfate sodium (PPS) is the active ingredient in ELMIRON, a drug approved for the relief of bladder pain associated with interstitial cystitis. The study objective was to characterize the pharmacokinetic and metabolic profiles of PPS following oral dosing of [3H]PPS. As specific assays for PPS do not exist, metabolic profiling was accomplished through multiple fraction collections and radiochromatographic techniques. Two groups of eight healthy female subjects sequentially received a single oral dose of 200 microCi [3H]PPS supplemented with 300 mg unlabelled PPS or 300 microCi [3H]PPS supplemented with 450 mg unlabelled PPS. Most of the administered dose (84%) was excreted in faeces as intact PPS, and a smaller percentage (6%) was excreted in urine. In summary, orally administered PPS was very poorly absorbed, with the majority of the drug being excreted in faeces as intact PPS and in urine as low molecular weight and desulfated PPS.

Level of functioning, severity of illness, and smoking status among chronic psychiatric patients.

It was hypothesized that chronic psychiatric patients who had quit smoking would be more functional and have lower Brief Psychiatric Rating Scale (BPRS) scores than those who continued to smoke. We interviewed 300 chronic psychiatric patients followed in the community. Fourteen percent were former smokers and nearly 11% had never smoked. Fifty-six percent of the sample were current smokers who had no intention of quitting, 13% were considering quitting, and 6% were seriously preparing to quit or had actually quit for a short period. When compared with current smokers, former smokers were more likely to live independently (p < .026) and less likely to have a drug or alcohol problem (p < .013). A random sample of current smokers were compared with former smokers on the BPRS. Former smokers had lower total BPRS scores (p < .03), and lower withdrawal/retardation subscale scores (p < .0058) than current smokers. We concluded that better functioning patients who smoked would be more likely to quit.

Metabolic activation of tris(2,3-dibromopropyl)phosphate to reactive intermediates. I. Covalent binding and reactive metabolite formation in vitro.

Analogs of tris(2,3-dibromopropyl)phosphate (Tris-BP) either labeled at specific positions with carbon-14, phosphorus-32, or oxygen-18 or dual-labeled with both deuterium and tritium were used as metabolic probes to study the chemical and metabolic events in the bioactivation of Tris-BP to chemically reactive metabolites in liver microsomal preparations. Oxidation at the terminal (C-3) carbon atom of the propyl groups of Tris-BP yielded the direct-acting mutagen 2-bromoacrolein as the major metabolite that binds to DNA. Although this reactive metabolite also appears to bind to microsomal protein, the rate of binding of radiolabeled Tris-BP to protein is 15-20x greater than binding to DNA, and some metabolites that retain the phosphate group are bound. Studies with deuterated analogs of Tris-BP implicate oxidation at C-2 of the propyl group as a major pathway that leads to protein binding which is enhanced by phenobarbital pretreatment of rats. Moreover, investigations with 18O-Tris-BP and H2(18)O show that Bis-BP that is formed from oxidation of Tris-BP incorporates one atom of oxygen from water. Deuterium isotope studies suggest that most of the Bis-BP arises from initial oxidation at C-2. Taken together these studies indicate that P-450 oxidation of Tris-BP at C-2 of the propyl group yields a reactive alpha-bromoketone metabolite of Tris-BP that can either alkylate proteins directly or be hydrolyzed to Bis-BP and an alpha-bromo-alpha'-hydroxyketone that can alkylate microsomal proteins.

Metabolic activation of tris(2,3-dibromopropyl)phosphate to reactive intermediates. II. Covalent binding, reactive metabolite formation, and differential metabolite-specific DNA damage in vivo.

Analogs of tris(2,3-dibromopropyl)phosphate (Tris-BP) either labeled at specific positions with carbon-14 and phosphorus-32 or dual-labeled with both deuterium and tritium were administered to male Wistar rats at a nephrotoxic dose of 360 mumol/kg. The covalent binding of Tris-BP metabolites to hepatic, renal, and testicular proteins was determined after 9 and 24 hr, and plasma concentrations of bis(2,3-dibromopropyl)-phosphate (Bis-BP) formed metabolically from Tris-BP were measured at intervals throughout the initial 9-hr postdosing period. The covalent binding of 14C-Tris-BP metabolites in the kidney (2495 +/- 404 pmol/mg protein) was greater than that in the liver (476 +/- 123 pmol/mg protein) or testes (94 +/- 11 pmol/mg protein); the extent of renal covalent protein binding of Tris-BP metabolites was decreased by 82 and 84% when deuterium was substituted at carbon-2 and carbon-3, respectively. Substitution of Tris-BP with deuterium at carbon-2 or carbon-3 also decreased the mean area under the curve for Bis-BP plasma concentration by 48 and 57%, respectively. The mechanism of Tris-BP-induced renal and hepatic DNA damage was evaluated in Wistar rats by an automated alkaline elution procedure after the administration of analogs of Tris-BP or Bis-BP labeled at specific positions with deuterium. Renal DNA damage was decreased when Tris-BP was substituted with deuterium at either carbon-2 or carbon-3; the magnitude of the change correlated with both a decrease in the area under the Bis-BP plasma curve and a decrease in renal covalent binding of Tris-BP metabolites for each of the deuterated analogs. In marked contrast, analogs of Bis-BP labeled with deuterium at carbon-2 or carbon-3 did not show a decrease in the severity of renal DNA damage compared to unlabeled Bis-BP. On the basis of these observations a metabolic scheme for hepatic P-450-mediated oxidation at either carbon-2 or carbon-3 of Tris-BP affording Bis-BP by two alternate pathways that are susceptible to primary deuterium kinetic isotope effects is proposed. The Tris-BP metabolite, Bis-BP, is subsequently metabolized to reactive intermediates that cause DNA damage and bind to kidney proteins in a mechanism independent of cytochrome P-450.

Investigations of mechanisms of reactive metabolite formation from (R)-(+)-pulegone.

1. (R)-(+)-Pulegone is a monoterpene that is oxidized by cytochromes P-450 to reactive metabolites that initiate events in the pathogenesis of hepatotoxicity in mice, rats and humans. 2. Selective labelling of (R)-(+)-pulegone with deuterium revealed that menthofuran was a proximate hepatotoxic metabolite formed by oxidation of the allylic methyl groups of pulegone. Incubations of pulegone with mouse liver microsomes in an atmosphere of 18O2 resulted in the formation of menthofuran that contained only oxygen-18 in the furan moiety. These results are consistent with oxidation of pulegone to an allylic alcohol that reacts intramolecularly with the ketone moiety to form a hemiketal that subsequently dehydrates to generate menthofuran. 3. Studies on the metabolism of menthofuran revealed that it is oxidized by cytochromes P-450 to an electrophilic gamma-ketoenal that reacts with nucleophilic groups on proteins to form covalent adducts. In addition, diastereomeric mintlactones are formed. Investigations with H2(18)O and 18O2 are indicative of a furan epoxide intermediate, or a precursor, in the formation of the gamma-ketoenal and mintlactones.

Reactive intermediates in the oxidation of menthofuran by cytochromes P-450.

Menthofuran, a naturally occurring hepatotoxin, is metabolically activated to chemically reactive intermediates that are capable of covalent binding to cellular proteins. Studies in vivo and in vitro with inhibitors and inducers of hepatic cytochromes P-450 demonstrated an association between hepatocellular damage caused by menthofuran and its metabolic activation and covalent binding to target organ proteins. The same gamma-ketoenal formed from the metabolic precursor of menthofuran, pulegone, is the major electrophilic metabolite of menthofuran as well. Diastereomeric mintlactones also are formed, and studies with H218O and 18O2 indicate that the gamma-ketoenal is a precursor to the mintlactones, as well as other reactive intermediates in the cytochrome P-450 mediated oxidation of menthofuran.

Metabolism of alachlor by rat and mouse liver and nasal turbinate tissues.

The metabolism of alachlor was studied using in vitro incubations with microsomal fractions prepared from liver and nasal turbinates of rats and mice. Specifically, the transformation of alachlor to 3,5-diethylbenzoquinone-4-imine was examined. A key intermediate in this pathway was identified as 2,6-diethylaniline, the formation of which required catalysis by microsomal arylamidases. 2,6-Diethylaniline was oxidized to 4-amino-3,5-diethylphenol and the electrophilic 3,5-diethylbenzoquinone 4-imine. Rat nasal tissue possessed high enzymatic activity which can promote the formation of the reactive quinone imine. Whole body autoradiographic analysis demonstrated localization of radioactivity in the rat nasal tissue following oral administration of alachlor. A methylsulfide metabolite of alachlor was shown to be a precursor to 2,6-diethylaniline. The deposition of radioactivity in the rat nasal tissue was more pronounced following oral administration of the methylsulfide metabolite of alachlor.

Metabolic activation of (R)-(+)-pulegone to a reactive enonal that covalently binds to mouse liver proteins.

Pulegone, a naturally occurring hepatotoxin, is metabolically activated to chemically reactive intermediates that are capable of covalent binding to cellular protein. Studies in vivo and in vitro with inhibitors and inducers of cytochrome P-450 demonstrated an association among the hepatocellular toxicity of pulegone and its metabolic activation and covalent binding to protein. The exocyclic double bond of pulegone apparently is an important structural feature in the activation mechanism and binding to protein inasmuch as the reduced analogue, menthone, is neither hepatotoxic nor does it bind extensively to tissue proteins. Preliminary studies using semicarbazide as a trapping agent indicate that an unsaturated gamma-ketoaldehyde is the ultimate chemically reactive metabolite of pulegone.

The metabolism of the abortifacient terpene, (R)-(+)-pulegone, to a proximate toxin, menthofuran.

(R)-(+)-Pulegone, the major monoterpene component of the abortifacient mint oil, pennyroyal oil, is metabolized by hepatic microsomal monooxygenases of the mouse to a hepatotoxin. The formation of a toxic metabolite is apparently mediated by cytochromes P-450 of the phenobarbital class inasmuch as phenobarbital pretreatment of mice increases, whereas beta-naphthoflavone pretreatment decreases, the extent of hepatic necrosis caused by pulegone. Furthermore, two inhibitors of cytochromes P-450, cobaltous chloride and piperonyl butoxide, block toxicity. An analog of (R)-(+)-pulegone that was labeled with deuterium in the allylic methyl groups was found to be significantly less hepatotoxic than the parent compound. The results indicate that oxidation of an allylic methyl group is required for generation of a hepatotoxic metabolite. Menthofuran was identified as a proximate toxic metabolite of (R)-(+)-pulegone, and investigations with (R)-(+)-pulegone-d6 and 18O2 strongly indicate that menthofuran is formed by a sequence of reactions that involve: 1) oxidation of an allylic methyl group, 2) intramolecular cyclization to form a hemiketal, and 3) dehydration to form the furan.

Biotransformation of olivetol by Syncephalastrum racemosum.

A study of the biotransformation of olivetol by Syncephalastrum racemosum ATCC 18192 has led to the isolation of three metabolites, which were identified as 4'-hydroxy-olivetol, 3-(3,5-dihydroxyphenyl)-1-propanol, and 3-(3,5-dihydroxyphenyl)-1-propanoic acid. The structures of the isolated metabolites were deduced by comparison of their spectral properties (pmr, cmr, ms) with those of olivetol. The absolute configuration of 4'-hydroxy-olivetol was determined to be R by the Horeau partial resolution method. Biotransformation of olivetol therefore appears to occur by a subterminal oxidation process.