Comparative in vitro activity of 16 antimicrobial agents against Actinobacillus pleuropneumoniae.

Sixteen antimicrobial agents were tested for their activity against 68 isolates of Actinobacillus pleuropneumoniae by determining the minimum inhibitory concentrations (MICs). Ceftiofur and the fluoroquinolones danofloxacin and enrofloxacin were the most active compounds, with a MIC for 90% of the isolates (MIC90) of (0.05 microg/ml. The MIC90 values of benzylpenicillin, amoxicillin and aspoxicillin were 0.78 units/ml, 0.39 microg/ml and < or = 0.05 microg/ml, respectively. Three isolates (4.4%) were resistant to penicillins, but aspoxicillin was as active as ceftiofur against the susceptible isolates, with MICs of < or = 0.05 microg/ml for all isolates. Resistance to oxytetracycline, chloramphenicol and thiamphenicol occurred in 22 (32.4%), 14 (20.6%) and 15 (22.1%) of the isolates, respectively. Doxycycline was more active than oxytetracycline, with a MIC90 of 1.56 microg/ml as against 25 microg/ml. Florfenicol was not only as active as thiamphenicol, with a MIC for 50% of the isolates (MIC50) of 0.39 microg/ml, but also active against thiamphenicol-resistant isolates. All the isolates were susceptible to florfenicol. All the isolates were also susceptible to gentamicin, spectinomycin, tilmicosin, colistin and tiamulin. Of these, spectinomycin was the least active, with a MIC50 of 25 microg/ml, followed by tiamulin, with a MIC50 of 6.25 microg/ml. Of the 68 isolates tested, 49 (72.0%) were of serotype 2; 14 (20.5%) were of serotype 1; 2 each (3.0%) were of serotypes 5 and 6; and one was of serotype 7. Of the isolates, 23 (33.8%) were resistant to one or more of the major antibiotics. Antibiotic resistance was found only infrequently among serotype 2, with 5 (10.2%) of 49 isolates being resistant to chloramphenicol and/or oxytetracycline, while it occurred in 18 (94.7%) of the 19 isolates of other serotypes.

Antimicrobial susceptibility of Pasteurella multocida isolated from cattle and pigs.

Minimum inhibitory concentrations (MICs) of 10 antimicrobial agents were determined for Pasteurella multocida from cattle and pigs (72 and 68 isolates, respectively). Higher MICs were observed with oxytetracycline, doxycycline, tilmicosin and thiamphenicol for porcine isolates than for bovine isolates. Enrofloxacin was the most active, with an MIC for 90% of the isolates (MIC90) of 0.05 microg/ml for both bovine and porcine isolates. Aspoxicillin exhibited the same excellent activity against penicillin-susceptible isolates as ceftiofur, with MICs ranging from < or = 0.025 to 0.1 microg/ml. Aminoglycosides were less active, with an MIC90 of > 100 microg/ml for both bovine and porcine isolates.

Antimicrobial susceptibilities of enterococci isolated from faeces of broiler and layer chickens.

Enterococci were isolated from faecal droppings of chickens in broiler and layer farms and the susceptibilities to nine therapeutic antimicrobial agents and six growth-promoting antibiotics were determined by the agar dilution method. Resistance to therapeutic antimicrobial agents such as ampicillin, clindamycin, erythromycin, streptomycin, tetracycline or tylosin was more frequent in enterococcal isolates from broiler farms than in those from layer farms. Resistance to ofloxacin was rare, occurring in only one (0.7%) of the Enterococcus faecium isolates from broiler farms. Resistance to growth-promoting antibiotics such as avilamycin, salinomycin and virginiamycin was common among isolates from broiler farms. Of the E. faecium isolates from broiler farms, 12.4% were resistant to avilamycin and 27.4% were resistant to virginiamycin. Resistance to salinomycin was detected in all enterococcal species, ranging from 12.4% of E. faecium isolates to 50% of E. hirae isolates.

Influence of antibiotics used as feed additives on the immune effect of erysipelas live vaccine in swine.

To investigate the influence of antibiotics used as feed additives on the immune response to erysipelas live vaccine, the pig inoculation test was applied. Avilamycin, oxytetracycline quaternary salt, enramycin, virginiamycin and tylosin phosphate were selected as test antibiotics. Five experimental feeds containing each antibiotic at the highest concentration permitted for feed additives in Japan, and the basal diet lacking antibiotics were examined. Twenty-nine pigs were divided into six groups. At first all the groups were fed with the antibiotic-free basal diet for 7 days, and then each group received the experimental feeds. On the 14th day after feeding with test feeds all the pigs, except for one control pig in each group, were immunized with the vaccine and all the pigs were then challenge-exposed to a virulent strain of Erysipelothrix rhusiopathiae 14 days after vaccination. The clinical response was observed every day for 14 days. In all the groups, most of the vaccinated pigs did not develop any clinical signs of acute erysipelas after the challenge exposure, whereas non-vaccinated control pigs died or showed severe generalized erythema with profound depression and anorexia. No differences in the protection against the challenge exposure were observed among the groups. Therefore, the present results suggest that these selected antibiotics would not interfere with the immune effect of the vaccine if given at the usual concentrations used for feed additives.

Use of ELISA for in vitro potency test of rabies vaccines for animal use.

The use of ELISA for the potency test of inactivated rabies vaccines for animal use was evaluated. Sandwich ELISA was developed, using monoclonal antibodies which enabled to recognize the glycoprotein (G-protein) of rabies virus and showed a protective effect in mice. Soluble G-protein which displays an ELISA activity but a low immunogenicity was removed by gel filtration before ELISA was carried out. The ELISA titres of the first fraction of vaccines separated by gel filtration were correlated with the neutralizing antibody titres of vaccinated guinea-pigs. Our results suggest that the ELISA titres of the first fraction of vaccines separated by gel filtration reflected the protective G-protein contents for the in vitro potency test of rabies vaccines compared with the current in vivo test.

Enzyme immunoassay of sulfamethoxazole in chicken tissues: interlaboratory study.

Three laboratories participated in the interlaboratory study of an enzyme immunoassay (EIA) method for determination of sulfamethoxazole (SMX) using samples of the same muscle, liver and plasma from chickens administered with SMX. Interlaboratory variation in the determined values were similar to those of other studies determined by gas chromatography and liquid chromatography. It was suggested that the interlaboratory difference in determined SMX residues from chicken tissues did not become problems of using EIA method, in spite of the differences of equipments and operators.

Cellular fatty acid composition of Erysipelothrix rhusiopathiae and Erysipelothrix tonsillarum.

The cellular fatty acid compositions in 6 strains of Erysipelothrix rhusiopathiae and 7 strains of Erysipelothrix tonsillarum were determined by gas chromatography. Fatty acids, ranging from C10 to C18, were detected in the test strains. The fatty acid profile was characterized by very high percentages of 18:1 (cis-9) (cis-9-octadecenoic acid; 72.4 to 82.1%) and 16:1 (hexadecenoic acid; 8.7 to 13.7%). The profiles of the E. rhusiopathiae and E. tonsillarum strains resembled each other, indicating that discrimination between E. rhusiopathiae and E. tonsillarum from qualitative or quantitative fatty acid differences is difficult.

The pharmacokinetics of primaquine in calves after subcutaneous and intravenous administration.

The pharmacokinetics of primaquine was studied in calves of 180-300 kg live weight. Primaquine was injected at 0.29 mg/kg (0.51 mg/kg as primaquine diphosphate) intravenously (IV) or subcutaneously (SC) and the plasma concentrations of primaquine and its metabolite carboxyprimaquine were determined by high-performance liquid chromatography. The extrapolated concentration of primaquine at zero time after IV administration was 0.50 +/- 0.48 microgram/ml (mean +/- SD) which decreased with an elimination half-life of 0.16 +/- 0.07 h. Primaquine was rapidly converted to carboxyprimaquine after either route of administration. The peak concentration of carboxyprimaquine was 0.50 +/- 0.08 microgram/ml at 1.67 +/- 0.15 h after IV administration. The corresponding value was 0.47 +/- 0.07 micrograms/ml at 5.05 +/- 1.20 h after SC administration. The elimination half-lives of carboxyprimaquine after IV and SC administration were 15.06 +/- 0.99 and 12.26 +/- 3.06 h, respectively. The areas under the concentration-time curve for carboxyprimaquine were similar following either IV or SC administration of primaquine; the values were 11.85 +/- 2.62 micrograms.h/ml after the former and 10.95 +/- 2.65 micrograms.h/ml after the latter. The mean area under the concentration-time curve for primaquine was less than 0.1 micrograms.h/ml after either route of administration.

High-performance liquid chromatographic determination of pamaquine, primaquine and carboxy primaquine in calf plasma using electrochemical detection.

A high-performance liquid chromatographic method with electrochemical detection is described for quantification of pamaquine, primaquine and carboxy primaquine in calf plasma. After the proteins had been precipitated with acetonitrile, the drugs were separated on a 5-microns C18-modified polymer gel column with an isocratic mobile phase. The detection limit was 0.01 microgram/ml in plasma for all three compounds. The applicability of the method in pharmacokinetic studies was demonstrated by determining the plasma concentrations of the three substances in calves administered a single dose of pamaquine or primaquine.

Pharmacokinetic data of bromofenofos in calves.

Bromofenofos (BF) was administered orally to six calves at a dose of 12 mg/kg and the concentrations of BF and its metabolite dephosphate bromofenofos (DBF) in plasma were determined using a high-performance liquid chromatographic method. DBF reached maximum concentration of 45.9 +/- 16.3 micrograms/ml, which occurred at 1.3 +/- 0.4 days. The elimination half life was calculated to be 1.8 +/- 0.3 days and the area under the plasma concentration versus time curve was 185.1 +/- 49.4 micrograms.day/ml. The ultrafiltrate of plasma samples revealed no free metabolite, indicating a binding of greater than 99% to plasma protein. DBF disappeared from plasma in all calves 21 days after administration. No BF was detected in plasma at any time.

Plasma concentration and placental transfer of bromofenofos in rats.

Bromofenofos (BF) was administered by gavage once to non-pregnant and pregnant rats at 50 mg/kg and the plasma concentration-time curves of BF and its metabolite dephosphate bromofenofos (DBF) were determined by high-performance liquid chromatography. DBF reached a peak at 9 h, with 113.4 +/- 5.2 micrograms/ml, followed by a subsequent decline with a plasma half-life of approximately 24 h, but BF was not detected at any time. Next, the concentration of DBF in the conceptus was determined in rats administered BF (50 mg/kg) on day 10 of pregnancy. The concentration in the conceptus did not exceed 30% of that in the maternal plasma at all times, with a maximum of 32.3 +/- 3.3 micrograms/g at 12 h. No BF was detected in maternal plasma. Third, BF (50 mg/kg) was administered to rats on day 15 of pregnancy to determine whether DBF could cross the placenta. The concentration of DBF in the placenta was nearly half that in the maternal plasma at all times. The fetal concentration was approximately 20% of the concentration in the maternal plasma by 24 h. DBF was detected in the amniotic fluid at all times, but the concentration was low, with a maximum of 11.1 +/- 1.4 micrograms/ml at 12 h.

Liquid chromatographic determination of sulfamoyldapsone in swine tissues.

A liquid chromatographic (LC) method is described for the quantitative determination of sulfamoyldapsone (2-sulfamoyl-4,4'-diaminodiphenyl sulfone) in swine muscle, liver, kidney, and fat. Sulfamoyldapsone was extracted from tissues with acetonitrile saturated with n-hexane. The extract was washed with n-hexane saturated with acetonitrile, concentrated, and cleaned up by alumina column chromatography. Sulfamoyldapsone was separated on an ODS column by using acetonitrile-methanol-water (6 + 18 + 76) and was detected at 292 nm. Overall average recovery of sulfamoyldapsone added to tissues at levels of 0.1 and 0.5 microgram/g was 93.3% +/- 6.0. Detection limit was 0.02 microgram/g in these tissues.

Cytotoxic lymphocytes in rat tumor in situ: effect of intraperitoneal injections of Propionibacterium avidum.

In vitro cytotoxicity against tumor cells of lymphocytes in sc implanted BC47 bladder tumor of ACI/N rats with or without Propionibacterium avidum (P. avidum) treatment was studied. Tumor-associated lymphoid cells (TAL) were obtained from tumor tissues by mechanical treatment (NDi fraction) and by enzymatic treatment with Dispase I, a proteolytic enzyme (Di fraction), followed by passage through glass wool columns to deplete tumor cells. NDi fraction of TAL from P. avidum-treated animals showed a significant cytolytic activity against BC47 cells, but not against other ACI/N bladder tumor cell lines, BC12 and BC50. These TAL lost the cytolytic activity on treatment with anti-rat thymocyte serum or anti-rat T cell monoclonal antibodies, R1-3B3 and R1-10B5, and complement. Natural killer activity determined with YAC-1 cells was low in the cells of NDi fraction and scarcely detectable in the cells of Di fraction from both P. avidum-treated and untreated rats. These results indicate that the antigen-specific cytotoxic T cells in the tumor in situ are induced by in vivo P. avidum treatment. On the other hand, P. avidum treatment augmented nonspecific cytolytic activity of peripheral lymphoid cells such as plastic-nonadherent peritoneal cells, spleen cells and blood lymphocytes in normal and BC47-bearing rats. However, the antigen-specific cytolytic T cells were predominantly induced and recovered in the plastic nonadherent peritoneal cells of BC47-bearing rats by the treatment with P. avidum.