Norwegian patients and retail chicken meat share cephalosporin-resistant Escherichia coli and IncK/blaCMY-2 resistance plasmids.

OBJECTIVES: In 2012 and 2014 the Norwegian monitoring programme for antimicrobial resistance in the veterinary and food production sectors (NORM-VET) showed that 124 of a total of 406 samples (31%) of Norwegian retail chicken meat were contaminated with extended-spectrum cephalosporin-resistant Escherichia coli. The aim of this study was to compare selected cephalosporin-resistant E. coli from humans and poultry to determine their genetic relatedness based on whole genome sequencing (WGS). METHODS: Escherichia coli representing three prevalent cephalosporin-resistant multi-locus sequence types (STs) isolated from poultry (n=17) were selected from the NORM-VET strain collections. All strains carried an IncK plasmid with a blaCMY-2 gene. Clinical E. coli isolates (n=284) with AmpC-mediated resistance were collected at Norwegian microbiology laboratories from 2010 to 2014. PCR screening showed that 29 of the clinical isolates harboured both IncK and blaCMY-2. All IncK/blaCMY-2-positive isolates were analysed with WGS-based bioinformatics tools. RESULTS: Analysis of single nucleotide polymorphisms (SNP) in 2.5 Mbp of shared genome sequences showed close relationship, with fewer than 15 SNP differences between five clinical isolates from urinary tract infections (UTIs) and the ST38 isolates from poultry. Furthermore, all of the 29 clinical isolates harboured IncK/blaCMY-2 plasmid variants highly similar to the IncK/blaCMY-2 plasmid present in the poultry isolates. CONCLUSIONS: Our results provide support for the hypothesis that clonal transfer of cephalosporin-resistant E. coli from chicken meat to humans may occur, and may cause difficult-to-treat infections. Furthermore, these E. coli can be a source of AmpC-resistance plasmids for opportunistic pathogens in the human microbiota.

The role of whole genome sequencing in antimicrobial susceptibility testing of bacteria: report from the EUCAST Subcommittee.

Whole genome sequencing (WGS) offers the potential to predict antimicrobial susceptibility from a single assay. The European Committee on Antimicrobial Susceptibility Testing established a subcommittee to review the current development status of WGS for bacterial antimicrobial susceptibility testing (AST). The published evidence for using WGS as a tool to infer antimicrobial susceptibility accurately is currently either poor or non-existent and the evidence / knowledge base requires significant expansion. The primary comparators for assessing genotypic-phenotypic concordance from WGS data should be changed to epidemiological cut-off values in order to improve differentiation of wild-type from non-wild-type isolates (harbouring an acquired resistance). Clinical breakpoints should be a secondary comparator. This assessment will reveal whether genetic predictions could also be used to guide clinical decision making. Internationally agreed principles and quality control (QC) metrics will facilitate early harmonization of analytical approaches and interpretive criteria for WGS-based predictive AST. Only data sets that pass agreed QC metrics should be used in AST predictions. Minimum performance standards should exist and comparative accuracies across different WGS laboratories and processes should be measured. To facilitate comparisons, a single public database of all known resistance loci should be established, regularly updated and strictly curated using minimum standards for the inclusion of resistance loci. For most bacterial species the major limitations to widespread adoption for WGS-based AST in clinical laboratories remain the current high-cost and limited speed of inferring antimicrobial susceptibility from WGS data as well as the dependency on previous culture because analysis directly on specimens remains challenging. For most bacterial species there is currently insufficient evidence to support the use of WGS-inferred AST to guide clinical decision making. WGS-AST should be a funding priority if it is to become a rival to phenotypic AST. This report will be updated as the available evidence increases.

Large IncHI2-plasmids encode extended-spectrum ß-lactamases (ESBLs) in Enterobacter spp. bloodstream isolates, and support ESBL-transfer to Escherichia coli.

We investigated the prevalence of extended-spectrum ß-lactamases (ESBLs) in Enterobacter spp. bloodstream isolates from 19 hospital laboratories in Norway during 2011. A total of 62/230 (27%) isolates were resistant to third-generation cephalosporins and four (1.7%) were ESBL-positive; blaCTX -M-15 (n = 3) and blaSHV -12 (n = 1). This is comparable to the prevalence of ESBLs in clinical isolates of Escherichia coli and Klebsiella pneumoniae in Norway during the same period. All ESBL-positive isolates were multidrug resistant (MDR) and harboured plasmid-mediated quinolone resistance. Three isolates supported transfer of large IncHI2-plasmids harbouring ESBL- and MDR-encoding genes to E. coli recipients by in vitro conjugation.

Rapid evolution and spread of carbapenemases among Enterobacteriaceae in Europe.

Plasmid-acquired carbapenemases in Enterobacteriaceae, which were first discovered in Europe in the 1990s, are now increasingly being identified at an alarming rate. Although their hydrolysis spectrum may vary, they hydrolyse most ß-lactams, including carbapenems. They are mostly of the KPC, VIM, NDM and OXA-48 types. Their prevalence in Europe as reported in 2011 varies significantly from high (Greece and Italy) to low (Nordic countries). The types of carbapenemase vary among countries, partially depending on the cultural/population exchange relationship between the European countries and the possible reservoirs of each carbapenemase. Carbapenemase producers are mainly identified among Klebsiella pneumoniae and Escherichia coli, and still mostly in hospital settings and rarely in the community. Although important nosocomial outbreaks with carbapenemase-producing Enterobacteriaceae have been extensively reported, many new cases are still related to importation from a foreign country. Rapid identification of colonized or infected patients and screening of carriers is possible, and will probably be effective for prevention of a scenario of endemicity, as now reported for extended-spectrum ß-lactamase (mainly CTX-M) producers in all European countries.

Molecular characterization of VIM-producing Klebsiella pneumoniae from Scandinavia reveals genetic relatedness with international clonal complexes encoding transferable multidrug resistance.

VIM-producing Klebsiella pneumoniae (VPKP) has been identified as a source of hospital outbreaks and is prevalent particularly in the Mediterranean region. In this study we have characterized eight VPKP isolates identified in Scandinavia during 2005-2008. With the exception of one isolate, all were from patients with recent history of hospitalization abroad (Greece, n = 6; Turkey, n = 1). Multilocus sequence typing (MLST) resulted in five sequence types (STs), ST36 (n = 1), ST147 (n = 4), ST272 (n = 1), ST273 (n = 1) and ST383 (n = 1), which except for ST272 were part of putative international clonal complexes. All were multidrug resistant due to the presence of other resistance determinants, including extended-spectrum ß-lactamases (CTX-M-3, SHV-5 and SHV-12), 16S rRNA methylases (ArmA) and plasmid-mediated quinolone resistance determinants (QnrS). One isolate harboured a novel VIM-variant (VIM-26) while VIM-1 and VIM-19 were detected in six and one isolate, respectively. Two different genetic structures surrounding the bla(VIM) gene were identified in four isolates. In two isolates bla(VIM-1) and bla(VIM-26) were located in an integron similar to In-e541 (intI1;bla(VIM-1/-26);aacA7; dhfrI;aadA1;3'CS) while in the other two isolates bla(VIM-1) was located in an integron lacking 3'CS but with an IS26 element in the 3'end (intI1;bla(VIM-1);aac(6')-Ib;IS26), as identified in the IncN plasmid pKOX105. The bla(VIM) -genes were located on transferable plasmids ranging from ∼40 to ∼240 kb and associated with Tn21 in four isolates. PCR-based replicon typing indicated association of bla(VIM) with IncN (n = 3) and A/C (n = 1) broad-host-range plasmids but also with unknown replicons (n = 4). In conclusion, Scandinavian VPKP is associated with importation and genetically related to international clones encoding transferable plasmid-mediated multidrug resistance.

An evaluation of the duration of efficacy of emamectin benzoate in the control of Caligus curtus Müller infestations in Atlantic cod, Gadus morhua L.

The duration of efficacy of emamectin benzoate in the control of Caligus curtus infestations in Atlantic cod, Gadus morhua L., was studied following an administration of 50 µg kg⁻¹ for seven consecutive days. No lice were found on medicated fish when challenged 1 week (challenge 1) or 5 weeks (challenge 2) following termination of medication, whereas the mean abundance of lice among the unmedicated fish was 17.9 and 19.3 lice per fish in challenge 1 and 2, respectively. Muscle concentrations of 19.5 ± 8.2 ng g⁻¹ and 3.4 ± 0.9 ng g⁻¹, respectively, and skin concentrations of 23.1 ± 10.8 and 4.2 ± 1.0 ng g⁻¹, respectively, were found 27 and 55 days following the termination of medication. Tissue concentrations and the duration of efficacy indicate a dosing regime for emamectin in cod, similar to the regime used for Atlantic salmon, Salmo salar L.

A sensitive and specific phenotypic assay for detection of metallo-ß-lactamases and KPC in Klebsiella pneumoniae with the use of meropenem disks supplemented with aminophenylboronic acid, dipicolinic acid and cloxacillin.

Enterobacteriaceae producing carbapenemases, such as KPC or metallo-ß-lactamases (MBLs), have emerged on several continents. Phenotypic tests are urgently needed for their rapid and accurate detection. A novel carbapenemase detection test, comprising a meropenem disk, and meropenem disks supplemented with 730 µg of EDTA, 1000 µg of dipicolinic acid (DPA), 600 µg of aminophenylboronic acid (APBA), or 750 µg of cloxacillin, was evaluated against Klebsiella pneumoniae isolates with KPC (n = 34), VIM (n = 21), IMP (n = 4) or OXA-48 (n = 9) carbapenemases, and carbapenem-resistant Enterobacteriaceae with porin loss in combination with an extended-spectrum ß-lactamase (ESBL) (n = 9) or AmpC hyperproduction (n = 5). Commercially available diagnostics tablets from Rosco containing meropenem and the same inhibitors as described above (except EDTA) were also evaluated. An increased meropenem inhibition zone was sought in the presence of each added ß-lactamase inhibitor. APBA had excellent sensitivity for detecting K. pneumoniae with KPC enzymes. Isolates with combined AmpC hyperproduction and porin loss were also positive in the APBA test but, unlike KPC producers, showed cloxacillin synergy. Both DPA and EDTA had excellent sensitivity for detection of MBL-producing K. pneumoniae. However, EDTA showed poor specificity, with positive results noted for 1/9 ESBL-producing isolates, for 4/34 KPC-producing isolates, and for 4/9 OXA-48-producing isolates, whereas all of these were negative when DPA was used. The in-house test distinguished accurately between several different mechanisms mediating reduced susceptibility to carbapenems in Enterobacteriaceae. The commercial combination tablets from Rosco performed similarly to the in-house test, with the exception of one false-positive MBL result and one false-positive KPC result among the OXA-48 producers.

Comparison of disk diffusion, Etest and VITEK2 for detection of carbapenemase-producing Klebsiella pneumoniae with the EUCAST and CLSI breakpoint systems.

The aim of this study was to compare CLSI and EUCAST MIC and disk diffusion carbapenem breakpoints for the detection of carbapenemase-producing Klebsiella pneumoniae. K. pneumoniae strains with known KPC (n = 31) or VIM (n = 20) carbapenemases were characterized by disk diffusion (Oxoid) and Etest (bioMérieux) vs. imipenem, meropenem and ertapenem, and with VITEK2 (bioMérieux, five different cards). Extended-spectrum ß-lactamase (ESBL) testing was performed with VITEK2 (bioMérieux), ESBL combination disks (Becton Dickinson) and the ESBL Etest (bioMérieux). With CLSI and EUCAST MIC breakpoints, respectively, 11 and seven of the strains were susceptible to imipenem, 12 and eight to meropenem, and seven and none to ertapenem. The EUCAST epidemiological cut-off (ECOFF) values for meropenem and ertapenem identified all carbapenemase producers, whereas the imipenem ECOFF failed in five strains. All carbapenemase producers were detected with EUCAST disk diffusion breakpoints for ertapenem and meropenem, and four strains were susceptible to imipenem. CLSI disk diffusion breakpoints characterized 18 (imipenem), 14 (meropenem) and three (ertapenem) isolates as susceptible. When cards with a single carbapenem were used, detection failures with VITEK2 were four for imipenem, none for meropenem and one for ertapenem. Cards containing all three carbapenems had one to two failures. With ESBL combination disks, 21/31 KPC producers and 2/20 VIM producers were positive. With VITEK2, no VIM producers and between none and seven KPC producers were ESBL-positive. All carbapenemase producers were detected with the meropenem MIC ECOFF, or the clinical EUCAST breakpoint for ertapenem. EUCAST disk diffusion breakpoints for meropenem and ertapenem detected all carbapenemase producers. VITEK2 had between none and four failures in detecting carbapenemase producers, depending on the antibiotic card.

A single-dose pharmacokinetic study of emamectin benzoate in cod, Gadus morhua L., held in sea water at 9 degrees C.

The pharmacokinetic profile of the antiparasitic agent emamectin benzoate was studied in plasma after intravenous (i.v.) injection and in plasma, muscle and skin following oral (p.o.) administration to cod, Gadus morhua, held in sea water at 9 degrees C and weighing 100-200 g. Following i.v. injection, the plasma drug concentration-time profile showed two distinct phases. The plasma distribution half-life (t(1/2)alpha) was estimated as 2.5 h, the elimination half-life (t(1/2)beta) as 216 h, the total body clearance (Cl(T)) as 0.0059 L kg(-1) h(-1) and mean residence time (MRT) as 385 h. The volume of distribution at steady state, V(d(ss)), was calculated to be 1.839 L kg(-1). Following p.o. administration the peak plasma concentration (C(max)) was 15 ng mL(-1), the time to peak plasma concentration (T(max)) was 89 h and t(1/2)beta was 180 h. The highest concentration in muscle (21 ng g(-1)) was measured after 7 days and t(1/2)beta was calculated to be 247 h. For skin, a peak concentration of 28 ng g(-1) at 3 days was observed and a t(1/2)beta of 235 h was determined. The bioavailability following p.o. administration was calculated to be 38%.

Treating experimentally induced vibriosis (Listonella anguillarum) in cod, Gadus morhua L., with florfenicol.

This study was performed to determine the efficacy of orally administered florfenicol in the treatment of experimentally induced vibriosis (Listonella anguillarum) in cod, Gadus morhua. The L. anguillarum strain HI-610 was used. This strain has a minimal inhibitory concentration value of 0.5 mg L(-1) against florfenicol. Fifteen groups of 40 fish each were challenged by bath with 1.7 x 10(5) CFU mL(-1) for 1 h. Three days following challenge, medication with florfenicol was introduced in 12 of the groups. The dosages used were 10 mg kg(-1) day(-1) for 10 consecutive days in marine or salmonid pellets, 10 mg kg(-1) day(-1) for five consecutive days in marine pellets or administered at days 1, 2, 4, 6 and 8 following initiation of treatment. Among challenged unmedicated fish mortality started at day 3 post-challenge reaching a final cumulative mortality of 77% at day 15. The experiment was terminated at day 26. In the medicated groups, the majority of deaths occurred from days 3-7 post-challenge reaching final cumulative mortalities of 31% and 52%, respectively, for the fish given marine and salmonid pellets for 10 consecutive days. The fish treated with medicated marine pellets for five consecutive days and at days 1, 2, 4, 6 and 8 (sequential feeding) following initiation of treatment had cumulative mortalities of 52% and 38%, respectively. Survival of medicated fish in all groups was significantly (P < 0.005) greater than survival of challenged unmedicated fish. Furthermore, a significant difference (P < 0.001) in survival was found between fish treated for 10 consecutive days using marine pellets and the groups using marine pellets for five consecutive days and salmonid pellets for 10 consecutive days. Twenty four hours following last medication, six fish had mean plasma concentrations of 3.3 +/- 1.7 and 3.5 +/- 2.8 microg mL(-1), respectively, in fish treated for 10 consecutive days using marine and salmonid pellets. Corresponding values for fish treated for five consecutive days and by sequential feeding were 2.2 +/- 2.3 and 1.7 +/- 0.7 microg mL(-1), respectively.

A single-dose pharmacokinetic study of oxolinic acid and vetoquinol, an oxolinic acid ester, in cod, Gadus morhua L., held in sea water at 8 degrees C and in vitro antibacterial activity of oxolinic acid against Vibrio anguillarum strains isolated from diseased cod.

The pharmacokinetic properties of the antibacterial agent oxolinic acid and vetoquinol, the carbitol ester of oxolinic acid, were studied after intravenous (i.v.) and oral (p.o.) administration to 100-150 g cod, Gadus morhua L., held in sea water at 8 degrees C. Following i.v. injection, the plasma drug concentration-time profile showed two distinct phases. The distribution half-life (t1/2alpha) was estimated at 1.3 h, the elimination half-life (t1/2beta) as 84 h and the total body clearance (Cl(T)) as 0.047 L kg(-1) h(-1). The volume of distribution at steady state, Vd(ss) was calculated to be 5.5 L kg(-1), indicating good tissue penetration of oxolinic acid in cod. Following p.o. administration of oxolinic acid or vetoquinol, the peak plasma concentrations (C(max)) of oxolinic acid and the time to peak plasma concentrations (T(max) were estimated to be 1.2 and 2.5 microg mL(-1) and 24 and 12 h, respectively. The bioavailabilities of oxolinic acid following p.o. administration of oxolinic acid and vetoquinol were calculated to be 55 and 72%, respectively. The in vitro minimum inhibitory concentration (MIC) values of oxolinic acid against three strains of Vibrio anguillarum isolated from diseased cod were 0.016 microg mL(-1) (HI-610), 0.250 microg mL(-1) (HI-618) and 0.250 microg mL(-1) (HI-A21). Based on a MIC value of 0.016 microg mmL(-1) a single p.o. administration of 25 mg kg(-1) of oxolinic acid maintains plasma levels in excess of 0.064 microg mL(-1), corresponding to four times the MIC-value, for approximately 12 days. The analogous value for a single p.o. dose of 25 mg kg(-1) of oxolinic acid administered as vetoquinol was 13 days.

The efficacy of a single intraperitoneal injection of oxolinic acid in the treatment of bacterial infections in goldsinny wrasse (Ctenolabrus rupestris) and corkwing wrasse (Symphodus melops) studied under field and laboratory conditions.

The efficacy of a single intraperitoneal injection of oxolinic acid to control an outbreak of atypical Aeromonas salmonicida infection in goldsinny wrasse (Ctenolabrus rupestris) and in the treatment of systemic vibriosis in corkwing wrasse (Symphodus melops) was examined. In addition a field study was performed to examine the effect of medication on the survival rate of goldsinny wrasse in Atlantic salmon cages. Four groups of wild caught goldsinny wrasse, each of 50 fish, were treated with an intraperitoneal injection of propylene glycol:saline (50:50) (control) or 50 mg/kg oxolinic acid at a concentration of 50 mg/mL. Three days after medication the fish in all groups were treated by an intraperitoneal injection of prednisolone acetate and an increase in seawater temperature from 9.0 to 11.5 degrees C. Cumulative mortalities were 18% in the two groups treated with oxolinic acid and 94 and 100% in the unmedicated control groups, giving a 'relative percentage survival' (RPS) value of 82%. A laboratory maintained population of originally wild caught corkwing wrasse experiencing high daily mortality was treated with oxolinic acid (50 mg/kg) or propylene glycol:saline (control). Cumulative mortalities were 84% (control) and 42% (oxolinic acid medicated group) giving an RPS value of 50%. In a field investigation using goldsinny wrasse approximately 30% were medicated with oxolinic acid (50 mg/kg) prior to stocking in cages with Atlantic salmon. In two of three cages the cumulative mortality was significantly lower (P = 0.025 and P < 0.001) in the medicated groups.

Absorption, tissue distribution and excretion of flumequine and oxolinic acid in corkwing wrasse (Symphodus melops) following a single intraperitoneal injection or bath treatment.

The pharmacokinetic properties of the antibacterial agents oxolinic acid and flumequine were studied in corkwing wrasse (Symphodus melops) after either intraperitoneal injection or bath treatment. Following intraperitoneal administration the peak plasma concentrations (Cmax) and the time to peak plasma concentrations (Tmax) were estimated to be 2.0 microg/mL and 12 h, respectively, for oxolinic acid and 2.6 microg/mL and 12 h, respectively, for flumequine. In muscle, Cmax and Tmax were estimated to 6.7 microg/g and 12 h, respectively, for oxolinic acid with corresponding values of 8.5 microg/g and 13 h, respectively, for flumequine. In liver, Cmax and Tmax were calculated to 7.0 microg/g and 12 h, respectively, for oxolinic and 12.2 microg/g and 11 h, respectively, for flumequine. Elimination half-lives (t1/2 beta) of 26, 24 and 29 h, respectively, for plasma, muscle and liver were calculated for flumequine. For oxolinic acid two distinct elimination phases were found and calculated to be 16 h (t1/2 beta) and 57 h (t1/2 gamma) in plasma, 15 and 59 h, respectively, in muscle and 20 and 72 h, respectively, in liver. Bath treatment using 150 mg/L of flumequine or 200 mg/L of oxolinic acid for 72 h resulted in flumequine concentrations of 1.0 microg/mL in plasma, 5.0 microg/g in muscle and 12.4 microg/g in liver. Corresponding values for oxolinic acid were 1.0 microg/g in plasma, 2.5 microg/g in muscle and 4.9 microg/g in liver.

Diseases, prophylaxis and treatment of the Atlantic halibut Hippoglossus hippoglossus: a review.

After substantial investments in research, the Atlantic halibut Hippoglossus hippoglossus is now being cultivated commercially in Norway, Iceland, Scotland and Canada. As with other domesticated species, disease problems have been experienced. This review summarizes the current state of knowledge of diseases of the Atlantic halibut, and their diagnosis, prophylaxis and treatment. In economic terms, the most important losses have been suffered at the larval and juvenile stages. The most important infections are caused by nodaviruses, causative agents of Viral Encephalopathy and Retinopathy (VER), which are the major reason why Norway's production of halibut fry has been level since 1995. An aquatic birnavirus, Infectious Pancreatic Necrosis Virus, is also an important agent of mortality. Vibrio anguillarum, Flexibacter ovolyticus and atypical Aeromonas salmonicida are the major bacterial pathogens. The protozoan parasites recorded include Ichthyobodo sp., the microsporidium Enterocytozoon sp., Trichodina hippoglossi, and the metazoan pathogens include myxozoans, helminths, Entobdella hippoglossi, Lepeophtheirus hippoglossi and other parasitic copepods. Experimental vaccines have been tested against V anguillarum and atypical A. salmonicida, with good results. A recombinant vaccine against nodaviruses is under development. A few trials have been carried out on non-specific immunostimulants, but no such treatment is currently available. A number of efficacy and pharmacokinetic trials with various antibacterial agents have also been published.

Efficacy of orally administered oxolinic acid and Vetoquinol, an oxolinic acid ester, for the treatment of furunculosis in Atlantic salmon Salmo salar held in seawater.

This study was performed to determine the efficacy of orally administered oxolinic acid and Vetoquinol, an oxolinic acid ester, in the treatment of experimental induced furunculosis in Atlantic salmon Salmo salar held in seawater. Two strains of the causative bacterium Aeromonas salmonicida subsp. salmonicida, 1 sensitive (VI-88/09/03175) and 1 resistant (3475/90) to oxolinic acid, were used. In 2 trials, cohabitational challenges were performed by introducing 8 fish challenged in advance by an intraperitoneal injection of 2.2 x 10(4) colony forming units of strain 3475/90 (Trial 1) or strain VI-88/09/03175 (Trial 2) to 10 aquaria each containing 40 healthy fish. The treatment groups in both trials consisted of 4 groups receiving either oxolinic acid (2 groups) or Vetoquinol (2 groups) and 1 control group. An unchallenged, unmedicated group was used to determine the natural mortality in the population. The recommended therapeutic dose of 25 mg oxolinic acid kg-1 fish at Days 1, 2, 4, 6, 8 and 10 following initiation of treatment was used. Oral medication initiated at Day 10 (Trial 1) or Day 11 (Trial 2) following challenge significantly (p < 0.05) lowered the specific mortality in all drug-treated groups compared to the untreated control groups. Mortality in Vetoquinol-treated groups was significantly (p < 0.05) lower than in oxolinic acid-treated groups in Trial 1 whereas no significant (p < 0.05) difference in survival rate was found between the medicated groups in Trial 2.

Absorption, tissue distribution, metabolism and excretion of ormetoprim and sulphadimethoxine in Atlantic salmon (Salmo salar) after intravenous and oral administration of Romet.

1. Uptake, bioavailability, tissue disposition and elimination of sulphadimethoxine (SDM) and ormetoprim (OMP) were examined in Atlantic salmon (Salmo salar) following intravenous and oral administration of Romet at a dose of 5 mg OMP and 25 mg SDM kg-1 fish. 2. Plasma clearance was rapid for both drugs following a single i.v. dose, characterized by t1/2 alpha = 0.48 and 0.54h, t1/2 beta = 9.9 and 25.6h for SDM and OMP respectively with a volume of distribution (Vss) = 0.389 and 2.478 l kg-1. 3. Following oral administration, peak plasma concentrations of 1.13 and 9.99 micrograms ml-1 were achieved after 17.6 and 20.3h for OMP and SDM respectively. Bioavailabilities were 85% for OMP and 39% for SDM. 4. Oral administration revealed the highest concentration of OMP in kidney and liver whereas the highest concentrations of SDM were found in muscle and bile. 5. High concentrations of N4-acetylated SDM were found in the bile indicating significant metabolism of SDM.

Simultaneous determination of ormethoprim and sulphadimethoxine in plasma and muscle of Atlantic salmon (Salmo salar).

A rapid clean-up and high-performance liquid chromatographic method for the simultaneous determination of ormethoprim and sulphadimethoxine in plasma and muscle of Atlantic salmon (Salmo salar) has been developed. Sample preparation is based on protein precipitation using trichloroacetic acid or methanol for plasma and muscle, respectively. The drugs are separated using a reversed-phase C18 analytical column and phosphate buffer-acetonitrile (80:20, v/v) containing 1-heptanesodiumsulphonate and triethylamine, as mobile phase. Detection was performed at 270 nm. The average recovery of ormethoprim was 97.2% in muscle and 95.7% in plasma, whereas the average recovery of sulphadimethoxine was 86.5% in muscle and 90.2% in plasma. The limit of detection at a signal-to-noise ratio of 3 was 50 ng/g and 30 ng/ml for ormethoprim in muscle and plasma respectively and 30 ng/g and 15 ng/ml in muscle and plasma respectively for sulphadimethoxine.

High-performance liquid chromatographic determination of oxolinic acid residues in fish silage.

A sensitive high-performance liquid chromatographic procedure was developed for the determination of oxolinic acid in fish silage. Oxolinic acid was extracted with a mixture of McIlvaine buffer (pH 3.6) and methanol (55:45) and re-extracted into dichloromethane. After successive clean-up by liquid-liquid partitioning, oxolinic acid was determined by HPLC with fluorimetric detection (exitation at 325 nm, emission at 360 nm). The analytical column was 3-microns MOS-Hypersil (150 x 4.6 mm I.D.) and the mobile phase contained (A) 0.025 M oxalic acid (pH 3.2)-acetonitrile-methanol-tetrahydrofuran (80:2.5:15:2.5, v/v) and (B) oxalic acid (pH 3.2)-acetonitrile-methanol-tetrahydrofuran (50:20:25:5), v/v) with the following elution profile: 0-5 min, linear gradient from 50 to 100% B; 5-10 min, isocratic at 100% B; 10.1-15 min, isocratic at 50% A-50% B. The calibration graph was linear over the concentration range studied (0.025-0.2 microgram/g). The limit of detection was 0.01 microgram/g (signal-to-noise ratio = 4).