Toxicological and pathological review of concurrent occurrence of nitrite toxicity and Swine Fever in pigs.

BACKGROUND: Plant associated nitrate/nitrite poisoning in buffalo, cattle, goat and sheep had been reported from various parts of the world. Horses and pigs are considered less susceptible to nitrate poisoning. In this study epidemiology of rare outbreak of nitrate poisoning in combination with classical swine fever in a small pig farm was investigated for development of strategies to control and prevent such incidents in future. MATERIALS AND METHODS: Concurrent infection of nitrate toxicity and classical swine fever were recorded in district Nawanshahar, Punjab. Eight pigs suddenly fell sick and died 2 days after feeding barseem + oats and marriage waste food. Twelve pigs were sick exhibiting symptoms of anorexia, fever (104-105(o)F), depression, constipation followed by diarrhea, respiratory difficulty, tremors and staggering gait with recumbency in four completely off-feed pigs. Blotchy discolorations of the skin of extremities (ears and snout) were observed in three pigs. RESULTS: Hematological examination revealed marked leucopenia. Postmortem examination revealed dark brown colored blood evident on opening the carcass and presence of barseem, oats in stomach and intestines. Lymph nodes were swollen and hemorrhagic. Serosal surface of spleen show various infarcts and button ulcers were recorded in cecum and colon, pathognomic lesion of classical swine fever. Nitrate toxicity was confirmed on the basis of quantitative determination of nitrate in the biological material of sick and dead animals. Fodder samples were (barseem + oats) positive for diphenylamine blue (DPB) test, Nitrate concentration in offended barseem and oats were found to be 2612 ppm and 3344 ppm as nitrate nitrogen (No3-N), respectively. Excessive amount of nitrate in stomach contents (924-1365 ppm), liver (22-48 ppm) and kidney (17-22 ppm) of dead animals (n = 8) confirmed that death of pigs was due to toxicity induced by nitrate/nitrite. CONCLUSION: The green fodder should be used cautiously in pigs and screening of fodder with DPB test prior offering to animals is strongly recommended to contain the nitrate/nitrite toxicity risk.

Pharmacokinetics following intravenous administration and pharmacodynamics of cefquinome in buffalo calves.

Disposition following single intravenous injection (2 mg/kg) and pharmacodynamics of cefquinome were investigated in buffalo calves 6-8 months of age. Drug levels in plasma were estimated by high-performance liquid chromatography. The plasma concentration-time profile following intravenous administration was best described by a two-compartment open model. Rapid distribution of cefquinome was evident from the short distribution half-life (t ½ α = 0.36 ± 0.01 h), and small apparent volume of distribution (Vd area = 0.31 ± 0.008 L/kg) indicated limited drug distribution in buffalo calves. The values of area under plasma concentration-time curve, elimination half-life (t ½ ß ), total body clearance (ClB), and mean residence time were 32.9 ± 0.56 µg · h/mL, 3.56 ± 0.05 h, 60.9 ± 1.09 mL/h/kg, and 4.24 ± 0.09 h, respectively. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration of cefquinome were 0.035-0.07 and 0.05-0.09 µg/mL, respectively. A single intravenous injection of 2 mg/kg may be effective to maintain the MIC up to 12 h in buffalo calves against the pathogens for which cefquinome is indicated.

Population pharmacokinetics and pharmacokinetic/pharmacodynamic evaluation of marbofloxacin against Coagulase-negative staphylococci, Staphylococcus aureus and Mycoplasma agalactiae pathogens in goats.

Marbofloxacin is a broad-spectrum fluoroquinolone, and an extra-label use has been reported in horse, sheep and goat. However, extrapolation of dosage regimens from cattle to horse and small ruminants could lead to incorrect dosing due to pharmacokinetic differences among species, increasing the risk of antimicrobial resistance or toxicity. Pharmacokinetic properties of marbofloxacin, including PK/PD analysis, have been studied by intravenous, intramuscular and subcutaneous administration in lactating and non-lactating goats. A population pharmacokinetic model of marbofloxacin in goats was built using 10 pharmacokinetic studies after intravenous, intramuscular, and subcutaneous administration at a dose of 2, 5 and 10 mg/kg. Serum or plasma and milk concentration-time profiles were simultaneously fitted with a non-linear mixed effect model with Monolix software. Level of milk production (lactating and non-lactating) and health status (healthy and un-healthy) were retained as covariates on volume of distribution and clearance. Marbofloxacin concentrations were well described in plasma/serum and milk by the population model. Simulated dose regimens of marbofloxacin administered at 2, 5 and 10 mg/kg by intramuscular route for five days were evaluated (n = 5000 per group). Steady-state fAUCs for each dose regimen were obtained. Probability of target attainment of fAUC/MIC ratios were determined and PK/PDco values (highest MIC for which 90% of individuals can achieve a prior numerical value of the fAUC/MIC index) were established using Monte Carlo simulations (n = 50,000). MIC values for wild type isolates of Staphylococcus aureus, coagulase negative staphylococci, and Mycoplasma agalactiae were determined and tentative epidemiological cutoff (TECOFF) were obtained at 1.0, 0.5 and 0.5 mg/L, respectively. The PK/PDco for the dose regimen of 2 mg/kg/24 h and 5 mg/kg/24 h (0.125 and 0.25 mg/L) were lower than TECOFF (0.5 and 1 mg/L). The dosage regimen of 10 mg/kg/24 h was adequate for intermediate MIC values of 0.125-0.50 mg/L and could be effective for a population with a target fAUC/MIC ratio ˂ 48 for Coagulase negative staphylococci and Mycoplasma agalactiae, but not for Staphylococcus aureus. Results obtained in this study could be taken as a starting point by committees that set the clinical breakpoints and justifies expert rules to optimize marbofloxacin dose regimens.

Semi-Mechanistic Modeling of Florfenicol Time-Kill Curves and in silico Dose Fractionation for Calf Respiratory Pathogens.

An important application of time-kill curve (TKC) assays is determination of the nature of the best PK/PD index (fAUC/MIC or fT% > MIC) and its target value for predicting clinical efficacy in vivo. VetCAST (the veterinary subcommittee of EUCAST) herein presents semi-mechanistic TKC modeling for florfenicol, a long acting (96 h) veterinary antimicrobial drug licensed against calf pneumonia organisms (Pasteurella multocida and Mannheimia haemolytica) to support justification of its PK/PDbreakpoint and clinical breakpoint. Individual TKC assays were performed with 6 field strains of each pathogen (initial inoculum 107 CFU/mL with sampling at times at 0, 1, 2, 4, 8, and 24 h). Semi-mechanistic modeling (Phoenix NLME) allowed precise estimation of bacteria growth system (KGROWTH, natural growth rate; KDEATH, death rate; BMAX, maximum possible culture size) and florfenicol pharmacodynamic parameters (EMAX, efficacy additive to KDEATH; EC50, potency; Gamma, sensitivity). PK/PD simulations (using the present TKC model and parameters of a florfenicol population pharmacokinetic model) predicted the time-course of bacterial counts under different exposures. Of two licensed dosage regimens, 40 mg/kg administered once was predicted to be superior to 20 mg/kg administered at 48 h intervals. Furthermore, we performed in silico dose fractionation with doses 0 - 80 mg/kg administered in 1, 2 or 4 administrations over 96 h and for MICs of 0.5, 1, 2, 4 mg/L with 2 inoculum sizes 105 and 107 CFU/mL. Regression analysis (Imax model) demonstrated that i) fAUC/MIC outperformed fT% > MIC as PK/PD index and ii) maximum efficacy (IC90%) was obtained when the average free plasma concentration over 96 h was equal to 1.2 to 1.4 times the MIC of Pasteurella multocida and Mannheimia haemolytica, respectively.

VetCAST Method for Determination of the Pharmacokinetic-Pharmacodynamic Cut-Off Values of a Long-Acting Formulation of Florfenicol to Support Clinical Breakpoints for Florfenicol Antimicrobial Susceptibility Testing in Cattle.

The PK/PD cut-off (PK/PDCO) value of florfenicol for calf pathogens was determined for long acting formulations (MSD Nuflor® and a bioequivalent generic product). PK/PDCO is one of the three MICs considered by VetCAST, a sub-committee of the European Committee on Susceptibility Testing (EUCAST), to establish a Clinical Breakpoint for interpreting Antimicrobial Susceptibility Testing (AST). A population model was built by pooling three pharmacokinetic data sets, obtained from 50 richly sampled calves, receiving one of two formulations (the pioneer product and a generic formulation). A virtual population of 5,000 florfenicol disposition curves was generated by Monte Carlo Simulations (MCS) over the 96 h of the assumed duration of action of the formulations. From this population, the maximum predicted MIC, for which 90% of calves can achieve some a priori selected critical value for two PK/PD indices, AUC/MIC and T>MIC, was established. Numerical values were established for two bacterial species of the bovine respiratory disease (BRD) complex, Pasteurella multocida and Mannheimia haemolytica. It was concluded that the PK/PDCO of florfenicol for both AUC/MIC and T>MIC was 1 mg/L.

Pharmacokinetics, urinary excretion and plasma protein binding of danofloxacin following intravenous administration in buffalo calves (Bubalus bubalis).

Pharmacokinetics, urinary excretion and plasma protein binding of danofloxacin was investigated in buffalo calves following intravenous administration at the dose rate of 1.25 mg/kg to select the optimal dosage regimen of danofloxacin. Drug concentrations in plasma and urine were measured by microbiological assaying. In vitro plasma protein binding was determined employing the equilibrium dialysis technique. The distribution and elimination of danofloxacin were rapid, as indicated by values (mean +/-SD) of distribution half-life (t(1/2)alpha = 0.16 +/- 0.07 h) and elimination half-life (t(1/2)beta = 4.24 +/- 1.78 h), respectively. Volume of distribution at steady state (Vss) = 3.98 +/- 1.69 L/kg indicated large distribution of drug. The area under plasma drug concentration versus time curve (AUC) was 1.79 +/- 0.28 micrg/ml x h and MRT was 8.64 +/- 0.61 h. Urinary excretion of danofloxacin was 23% within 48 h of its administration. Mean plasma protein binding was 36% at concentrations ranging from 0.0125 microg/ml to 1 microg/ml. On the basis of pharmacokinetic parameters obtained, it is concluded that the revision of danofloxacin dosage regimen in buffalo calves is needed because the current dosage schedule (1.25 mg/kg) is likely to promote resistance.

Characterization of the oestrogenic activity of non-aromatic steroids: are there male-specific endogenous oestrogen receptor modulators?

BACKGROUND AND PURPOSE: The endogenous oestrogens have important biological functions in men as well as in women. Because 17beta-oestradiol and oestrone are also formed in the male body, these aromatic oestrogens are generally thought to be responsible for exerting the required oestrogenic functions in the male. In the present study, we tested the hypothesis that some of the non-aromatic steroids that are androgen precursors or metabolites with hydroxyl groups at C-3 and/or C-17 positions may also be able to serve as ligands for the oestrogen receptors (ER) in the male. EXPERIMENTAL APPROACH: A total of sixty non-aromatic steroids (selected from families of androstens, androstans, androstadiens, oestrens and oestrans) were analysed for their ability to bind and activate the human ERalpha and ERbetain vitro and in cultured cells. KEY RESULTS: Six of the non-aromatic steroids, that is, 5-androsten-3beta,17beta-diol, 5alpha-androstan-3beta,17beta-diol, 5(10)-oestren-3alpha,17beta-diol, 5(10)-oestren-3beta,17beta-diol, 4-oestren-3beta,17beta-diol and 5alpha-oestran-3beta,17beta-diol, were found to have physiologically relevant high binding affinity ( approximately 50% of that of oestrone) for human ERalpha and ERbeta. These non-aromatic steroids also activated the transcriptional activity of human ERs and elicited biological responses (such as growth stimulation) in two representative ER-positive human cancer cell lines (MCF-7 and LNCaP) with physiologically relevant potency and efficacy. Molecular docking analysis of these six active compounds showed that they could bind to ERalpha and ERbeta in a manner similar to that of 17beta-oestradiol. CONCLUSIONS AND IMPLICATIONS: These results provide evidence for the possibility that some of the endogenous androgen precursors or metabolites could serve as male-specific ER ligands.