Prevalence and Pathogenicity of Cryptosporidium suis in Pre‐ and Post‐weaned Pigs

A total of 4338 faecal samples, 135 of sows, 3368 of pre‐weaned and 835 of post‐weaned piglets from eight farms in South Bohemia, Czech Republic were collected and examined for Cryptosporidium infection. No sow, but 5.7% pre‐weaned and 24.1% post‐weaned piglets were positive for Cryptosporidium infection. No relationship was found between diarrhoea and Cryptosporidium infection in any of the different age groups (pre‐ and post‐weaned piglets). Four piglets, which were sporadically shedding cryptosporidia in faeces, were necropsied. Neither clinical signs of diarrhoea nor macroscopical changes were found. Histologically, a moderate infection of cryptosporidia was detected in the glandular epithelium along the large intestine, with predisposition to the ansa centralis of the colon. No inflammatory response in the lamina propria was observed. Cryptosporidia were also commonly found in the glandular epithelium of submucosal lymphoglandular complexes in the colon. Cryptosporidium isolates from all farms were identified as Cryptosporidium suis using molecular markers (SSU rRNA). All of the C. suis strains obtained were larger [6.2 (6.0–6.8) × 5.5 (5.3–5.7) μm] than any isolate described so far [4.6 (4.4–4.9) × 4.2 (4.0–4.3) μm] and did not appear to be infective for neonatal BALB/c mice.     

Genotypic Prevalence of the Adhesin Involved in Diffuse Adherence in Escherichia coli Isolates in Pre‐weaned Pigs with Diarrhoea in Korea

A total of 1002 Escherichia coli strains isolated from pre‐weaned pigs with diarrhoea on 1114 swine farms were screened for the presence of the adhesin involved in diffuse adherence (AIDA) gene by polymerase chain reaction (PCR). Escherichia coli isolates that carried AIDA genes were also tested by PCR for the detection of five fimbriae (F4, F5, F6, F18 and F41), heat‐stable (STa, STb) and heat‐labile (LT) enterotoxin, enteroaggregative E. coli heat‐stable enterotoxin 1 (EAST1), and Shiga toxin 2 oedema disease (Stx2e) genes. Twenty‐three (2.3%) of the 1002 E. coli isolates carried the gene for AIDA. Among 23 isolates shown to carry genes for AIDA, three carried the AIDA gene as the only shown virulence factor. Other isolates carried other virulence factor genes in addition to AIDA. Four isolates carried genes for at least one of the fimbrial adhesins and enterotoxins. Sixteen isolates carried genes for enterotoxins only. The AIDA may represent an additional virulence determinant in pre‐weaned pigs with diarrhoea.     

Antiviral Effect of Saccharomyces cerevisiaeβ‐glucan to Swine Influenza Virus by Increased Production of Interferon‐γ and Nitric Oxide

The aim of these experiments was to investigate the potential antiviral effect of Saccharomyces cerevisiaeβ‐glucan on the pneumonia induced by swine influenza virus (SIV). Forty colostrum‐deprived 5‐day‐old piglets were randomly divided into four groups of 10. The 20 pigs in groups 1 and 2 were administered Saccharomyces cerevisiaeβ‐glucan orally (50 mg/day/pig; En‐Bio Technology Co., Ltd) for 3 days before SIV infection and those in groups 3 and 4 were given culture medium/diluent alone. Groups 1 and 3 were inoculated intranasally with 3 ml of tissue culture fluid containing 2 × 10⁶ tissue culture infective doses 50% (TCID₅₀)/ml of SIV and those in groups 2 and 4 were exposed in the same manner to uninfected cell culture supernatant. The microscopic lung lesions induced by SIV infection (group 1 pigs) were significantly more severe than those induced by infection in animals pre‐administered β‐glucan (group 3) (P < 0.05). Significantly more SIV nucleic acid was detected in the lungs of pigs experimentally infected with SIV only (group 1) at 5, 7 and 10 days post‐inoculation (dpi) compared with lungs from pigs pre‐administered β‐glucan and infected with SIV (group 3) (P < 0.05). The concentrations of interferon‐γ (IFN‐γ) and nitric oxide (NO) in bronchoalveolar lavage fluid from pigs pre‐administered β‐glucan and infected with SIV (group 3) were significantly higher than for any other group at 7 and 10 dpi for IFN‐γ, and at 5, 7 and 10 dpi for NO (P < 0.05). Saccharomyces cerevisiaeβ‐glucan reduced the pulmonary lesion score and viral replication rate in SIV‐infected pigs. These findings support the potential application of β‐glucan as prophylactic/treatment agent in influenza virus infection.     

Effect of added dietary threonine on growth performance,health, immunity and gastrointestinal function of weaning pigs with differing genetic susceptibility to Escherichia coli infection and challenged with E. coli K88ac

Threonine (Thr) is important for mucin and immunoglobulin production. We studied the effect of added dietary Thr on growth performance, health, immunity and gastrointestinal function of weaning pigs with differing genetic susceptibility to E. coli K88ac (ETEC) infection and challenged with ETEC. Forty‐eight 24‐day‐old weaned pigs were divided into two groups by their ETEC susceptibility using mucin 4 (MUC4) gene as a marker (2 MUC4^?/?, not‐susceptible, and 2 MUC4^+/+, susceptible, pigs per litter). Within genotype, pigs were fed two different diets: 8.5 (LThr) or 9.0 (HThr) g Thr/kg. Pigs were orally challenged on day 7 after weaning and slaughtered on day 12 or 13 after weaning. Before ETEC challenge, HThr pigs ate more (p < 0.05). The diet did not affect post‐challenge growth, but HThr tended to increase post‐challenge feed efficiency (p = 0.087) and overall growth (p = 0.087) and feed efficiency (p = 0.055). Before challenge, HThr pigs excreted less E. coli (p < 0.05), while after challenge, diet did not affect the number of days with diarrhoea and ETEC excretion. MUC4^+/+ pigs responded to the challenge with more diarrhoea, ETEC excretion and anti‐K88 IgA in blood and jejunal secretion (p < 0.001). HThr pigs had a higher increase of anti‐K88 IgA values in jejunal secretion (p = 0.089) and in blood (p = 0.089, in MUC4^+/+ pigs only). Thr did not affect total IgA and IgM values, morphometry of jejunum, goblet cells count in colon, total mucin from jejunum and colon, but varied jejunal goblet cells counts (p < 0.05). In the first two post‐weaning weeks, 8.5 g Thr/kg diet may be not sufficient to optimize initial feed intake, overall feed efficiency and intestinal IgA secretion and to control the gut microbiota in the first post‐weaning week, irrespective of the pig genetic susceptibility to ETEC infection.     

Antibiotic resistant Escherichia coli in southeastern Australian pig herds and implications for surveillance

To investigate public health implications of antibiotics to control post‐weaning scours, we surveyed 22 commercial pig herds in southeastern Australia. Fifty faecal samples per herd were collected from pre‐ and post‐weaned piglets. Presumptive Escherichia coli isolates were confirmed by MALDI‐TOF MS. Isolates (n = 325) were screened for susceptibility to 19 veterinary antibiotics using MIC broth microdilution. All 325 E. coli isolates underwent further testing against 27 antibiotics used in human medicine and were screened for ETEC adhesin and enterotoxin genes (F4 (K88), F5 (K99), F6 (987P), F18, F41, STa, STb, Stx2e and LT) by multiplex PCR. Isolates identified as phenotypically resistant to third‐generation cephalosporin (3GC) and aminoglycoside antibiotics were screened by multiplex PCR/reverse line blot to detect common β‐lactam and aminoglycosides resistance genes, confirmed by sequencing. Twenty (6.1%) of the E. coli isolates were resistant to 3GC antibiotics and 24 (7.4%) to the aminoglycoside antibiotic gentamicin. Genetic analysis revealed six different extended spectrum β‐lactamase (ESBL) genes (blaCTX‐M‐1, ‐14, ‐15, ‐27, blaSHV‐12 and blaCMY‐2‐like genes), four of which have not been previously reported in Australian pigs. Critically, the prevalence of 3GC resistance was higher in non‐pathogenic (non‐ETEC) isolates and those from clinically normal (non‐diarrhoeal) samples. This highlights the importance of non‐ETECE. coli as reservoirs of antimicrobial resistance genes in piglet pens. Antimicrobial resistance surveillance in pig production focused on diagnostic specimens from clinically‐affected animals might be potentially misleading. We recommend that surveillance for emerging antimicrobial resistance such as to 3GC antibiotics should include clinically healthy pigs.     

Resistant cutoff values and optimal scheme establishments for florfenicol against Escherichia coli with PK‐PD modeling analysis in pigs

Florfenicol, a structural analog of thiamphenicol, has broad‐spectrum antibacterial activity against gram‐negative and gram‐positive bacteria. This study was conducted to investigate the epidemiological, pharmacokinetic–pharmacodynamic cutoff, and the optimal scheme of florfenicol against Escherichia coli (E. coli) with PK‐PD integrated model in the target infectious tissue. 220 E. coli strains were selected to detect the susceptibility to florfenicol, and a virulent strain P190, whose minimum inhibitory concentration (MIC) was similar to the MIC₅₀ (8 μg/ml), was analyzed for PD study in LB and ileum fluid. The MIC of P190 in the ileum fluid was 0.25 times lower than LB. The ratios of MBC/MIC were four both in the ileum and LB. The characteristics of time‐killing curves also coincided with the MBC determination. The recommended dosages (30 mg/kg·body weight) were orally administrated in healthy pigs, and both plasma and ileum fluid were collected for PK study. The main pharmacokinetics (PK) parameters including AUC_24 hr, AUC_0–∞, T_max, T_1/2, C_max, CL_b, and K_e were 49.83, 52.33 μg*h/ml, 1.32, 10.58 hr, 9.12 μg/ml, 0.50 L/hr*kg, 0.24 hr^?1 and 134.45, 138.71 μg*hr/ml, 2.05, 13.01 hr, 16.57 μg/ml, 0.18 L/hr*kg, 0.14 hr^?1 in the serum and ileum fluid, respectively. The optimum doses for bacteriostatic, bactericidal, and elimination activities were 29.81, 34.88, and 36.52 mg/kg for 50% target and 33.95, 39.79, and 42.55 mg/kg for 90% target, respectively. The final sensitive breakpoint was defined as 16 μg/ml. The current data presented provide the optimal regimens (39.79 mg/kg) and susceptible breakpoint (16 μg/ml) for clinical use, but these predicted data should be validated in the clinical practice.     

The Use of Tannins to Control Salmonella Typhimurium Infections in Pigs

The aim of this study was to determine whether a hydrolysable tannin extract of sweet chestnut wood (Globatan^®) has an inhibitory effect on Salmonella Typhimurium survival both in vitro and in vivo in pigs. In a first experiment, the minimal inhibitory concentration of Globatan^® on 57 Salmonella Typhimurium isolates was determined. For all isolates, an MIC of 160–320 μg/ml was found. The second in vitro study revealed that Salmonella growth was strongly reduced using Globatan^® concentrations of 25–50 μg/ml and nearly completely inhibited at a concentration of 100 μg/ml Globatan^®. In an in vivo trial, two groups of six piglets, each group receiving feed with or without the addition of Globatan^® (3 g/kg), were orally inoculated with 10⁷ colony forming units of a Salmonella Typhimurium strain. Globatan^® had no effect on faecal excretion of Salmonella, and no differences in colonization of the intestines and internal organs were demonstrated in pigs euthanized at 4 days post‐inoculation. In conclusion, the hydrolysable tannin extract used in this study showed strong action against Salmonella Typhimurium in vitro but not in vivo.     

Pharmacokinetics of marbofloxacin in pigs after intravenous and intramuscular administration of a single dose of 8 mg/kg: dose proportionality,influence of the age of the animals and urinary elimination

The pharmacokinetics of marbofloxacin in pigs were evaluated as a function of dose and animal age following intravenous and intramuscular administration of a 16% solution (Forcyl^®). The absolute bioavailability of marbofloxacin as well as the dose proportionality was evaluated in 27‐week‐old fattening pigs. Blood PK and urinary excretion of marbofloxacin were evaluated after a single intramuscular dose of 8 mg/kg in 16‐week‐old male pigs. An additional group of 12‐week‐old weaned piglets was used for the evaluation of age‐related kinetics. The plasma and urine concentration of marbofloxacin was determined using a HPLC method. Pharmacokinetic parameters were calculated using noncompartmental methods. After intravenous administration in 27‐week‐old fattening pigs, the total body clearance was 0.065 L/h·kg. After intramuscular administration to the same animals, the mean observed C_max was 6.30 μg/mL, and the AUC_INF was 115 μg·h/mL. The absolute bioavailability was 91.5%, and dose proportionality was shown within the dose range of 4–16 mg/kg. The renal clearance was about half of the value of the total clearance. The total systemic clearance values significantly decreased as a function of age, being 0.092 L/h·kg and 0.079 L/h·kg in pigs aged 12 and 16 weeks, respectively.     

A microbiological assay to estimate the antimicrobial activity of parenteral tildipirosin against foodborne pathogens and commensals in the colon of beef cattle and pigs

Tildipirosin (TIP) is a novel 16‐membered‐ring macrolide authorized for the treatment of bovine and swine respiratory disease. The pH dependency of macrolide antimicrobial activity is well known. Considering that the pH in the colon contents of growing beef cattle and pigs is usually below pH 7.0, the minimum inhibitory concentrations (MIC) of TIP against foodborne bacterial pathogens such as Campylobacter (C.) coli, C. jejuni and Salmonella enterica and commensal species including Enterococcus (E.) faecalis, E. faecium and Escherichia coli were determined under standard (pH 7.3 ± 1) or neutral as well as slightly acidic conditions. A decrease in pH from 7.3 to 6.7 resulted in an increase in MICs of TIP. Except for the MICs > 256 μg/mL observed in the resistant subpopulation of the C. coli and the Enterococcus species, the MIC ranges increased from 2–8 μg/mL to 64–> 256 μg/mL for Salmonella enterica and E. coli, from 8–16 μg/mL to 32–128 μg/mL for the two Campylobacter species, and from 4–32 μg/mL to 128–> 256 μg/mL for both Enterococcus species. To estimate the antimicrobial activity of TIP in the colon contents of livestock during recommended usage of the parenterally administered TIP (Zuprevo^®), and to compare this with the increased MICs at the slightly acidic colonic pH, we developed and validated a microbiological assay for TIP and used this to test incurred faecal samples collected from cattle and pigs. Microbiological activity of luminal TIP was determined in aqueous supernatants from diluted faeces, using standard curves produced from TIP‐spiked faecal supernatants. The limit of quantification (LOQ) for TIP was 1 μg/mL (ppm). In a cattle study (n = 14), 3 of 28 faecal samples collected 24 and 48 h post‐treatment were found to contain TIP above the LOQ (concentrations of 1.3–1.8 ppm). In another cattle study (n = 12) with faecal samples collected at 8, 24 and 48 h post‐treatment, TIP concentrations were above the LOQ in 4 of the 8 h samples (1.2–2.6 ppm) and one of the 24‐h samples (1.3 ppm). In a pig study (n = 12) with faecal samples collected 24, 48 and 72 h post‐treatment, only one sample contained TIP above the LOQ (concentration 1.5 ppm). In another pig study (n = 12), with samples collected at 8, 24 48 and 96 h post‐treatment, TIP concentrations were above the LOQ in one 8‐h sample (1.1 ppm) and two 24‐h samples (2.3 and 2.5 ppm). None of the 48‐h and 96‐h samples from these 4 studies contained measurable TIP concentrations. Thus, in cattle and pigs, only a small fraction of faecal samples collected up to 24 h postdosing contained measurable microbiologically active TIP, with its maximum limited to 2.6 μg/mL. This is several log₂ dilution steps below the MICs of TIP against foodborne pathogens and commensals collected under acidic conditions comparable with those in the colonic contents and may explain a lack of intestinal dysbacteriosis with parenteral tildipirosin in livestock.     

Pharmacokinetics of orbifloxacin in crucian carp (Carassius auratus) after intravenous and intramuscular administration

The pharmacokinetics of orbifloxacin was studied after a single dose (7.5 mg/kg) of intravenous or intramuscular administration to crucian carp (Carassius auratus ) reared in freshwater at 25°C. Plasma samples were collected from six fish per sampling point. Orbifloxacin concentrations were determined by high‐performance liquid chromatography with a 0.02 μg/ml limit of detection, then were subjected to noncompartmental analysis. After intravenous injection, initial concentration of 5.83 μg/ml, apparent elimination rate constant (λ_z) of 0.039 hr^?1, apparent elimination half‐life (T_1/2λz) of 17.90 hr, systemic total body clearance (Cl) of 75.47 ml hr^?1 kg^?1, volume of distribution (Vz) of 1,948.76 ml/kg, and volume of distribution at steady‐state (Vss) of 1,863.97 ml/kg were determined, respectively. While after intramuscular administration, the λ_z, T _1/2λz, mean absorption time (MAT ), absorption half‐life (T _1/2ka), and bioavailability were determined as 0.027 hr^?1, 25.69, 10.26, 7.11 hr, and 96.46%, respectively, while the peak concentration was observed as 3.11 ± 0.06 μg/ml at 2.0 hr. It was shown that orbifloxacin was completely but relatively slowly absorbed, extensively distributed, and slowly eliminated in crucian carp, and an orbifloxacin dosage of 10 mg/kg administered intravenously or intramuscularly would be expected to successfully treat crucian carp infected by strains with MIC values ≤0.5 μg/ml.     

Pharmacokinetics and pharmacodynamics of intravenous and transdermal flunixin meglumine in meat goats

The aim of this study was to determine the pharmacokinetics and prostaglandin E₂ (PGE₂) synthesis inhibiting effects of intravenous (IV) and transdermal (TD) flunixin meglumine in eight adult female Boer goats. A dose of 2.2 mg/kg was administered intravenously (IV) and 3.3 mg/kg administered TD using a cross‐over design. Plasma flunixin concentrations were measured by LC‐MS/MS. Prostaglandin E₂ concentrations were determined using a commercially available ELISA. Pharmacokinetic (PK) analysis was performed using noncompartmental methods. Plasma PGE₂ concentrations decreased after flunixin meglumine for both routes of administration. Mean λ_z‐HL after IV administration was 6.032 hr (range 4.735–9.244 hr) resulting from a mean V_z of 584.1 ml/kg (range, 357.1–1,092 ml/kg) and plasma clearance of 67.11 ml kg^?1 hr^?1 (range, 45.57–82.35 ml kg^?1 hr^?1). The mean C_max, T_max, and λ_z‐HL for flunixin following TD administration was 0.134 μg/ml (range, 0.050–0.188 μg/ml), 11.41 hr (range, 6.00–36.00 hr), and 43.12 hr (15.98–62.49 hr), respectively. The mean bioavailability for TD flunixin was calculated as 24.76%. The mean 80% inhibitory concentration (IC₈₀) of PGE₂ by flunixin meglumine was 0.28 μg/ml (range, 0.08–0.69 μg/ml) and was only achieved with IV formulation of flunixin in this study. The PK results support clinical studies to examine the efficacy of TD flunixin in goats. Determining the systemic effects of flunixin‐mediated PGE₂ suppression in goats is also warranted.     

Morphology of Dromedary Camel Oocytes and their Ability to Spontaneous and Chemical Parthenogenetic Activation

The present work was conducted to examine (1) the morphology of dromedary cumulus‐oocytes complexes (COCs), (2) to study the incidence of spontaneous development of oocytes in vivo and (3) to assess the ability of in vitro matured dromedary oocytes to chemical parthenogenetic activation compared with in vitro fertilized (IVF) oocytes. COCs were recovered from dromedary ovaries classified according to their morphology into six categories. Oocyte diameter was measured using eye piece micrometer. For chemical activation, COCs with at least three layers of cumulus‐cells were in vitro matured (IVM) in TCM 199 + 10 μg/ml FSH + 10 IU hCG/ml + 10% FCS + 50 μg/ml gentamycin. COCs were incubated for 40 h at 38.5°C under 5% CO₂ in humidified air. After IVM, matured oocytes with first polar body (first Pb) were divided into two groups. Group 1: activated in 7% ethanol (E) for 5 min followed by culture in 2 mM 6‐dimethylaminopurin (6‐DMAP, E D, subgroup 1) or 10 μg/ml cycloheximide (CHX, E CHX, subgroup 2) for 3.5 h at 38.5°C under 5% CO₂. In group 2, oocytes were activated using 50 μM Ca A23187 (Ca A) for 5 min followed by culture in 2 mM 6‐DMAP (Ca D, subgroup 3) or 10 μg/ml CHX(Ca CHX, subgroup 4) for 3.5 h at 38.5°C under 5% CO₂. For control group, IVM oocytes were fertilized using frozen‐thawed camel spermatozoa separated by swim‐up method then suspended in Fert‐TALP medium supplemented with 6 mg/ml BSA (FAF) + 10 μg/ml heparin. In all groups, oocytes were in vitro cultured in SOFaa medium + 5% FCS and 5 μg/ml insulin + 50 μg/ml gentamycin. Cleavage rate and embryo development were checked on Days 2, 5 and 8. An average of 11.3 ± 0.3 COCs were recovered/dromedary ovary. Categories 1 and 2 represented 33.1% and 34.8%, respectively, and were significantly higher (p < 0.01) than the other categories (19.1, 9.2 and 2.6% for categories 3–5, respectively). Category 6 (embryo‐like structures) represented 1.2% of the recovered oocytes, staining of these embryo‐like structures with orcien dye indicated the presence of divided cells with condensed nuclei. Dromedary oocytes averaged 166.2 ± 2.6 μm in diameter with black cytoplasm. Chemical activation of IVM dromedary oocyte with first Pb in 7% ethanol or 50 μM Ca A followed by culture in 2 mM 6‐DMAP showed significantly higher (p < 0.01) cleavage and developmental rates to the morula stage than oocytes activated using 7% ethanol or 50 μM Ca A followed by 10 μg/ml CHX or in vitro fertilized control group. Higher (p < 0.01) proportion of oocytes sequentially cultured in 10 μg/ml CHX or that in vitro fertilized were arrested at the 2–4‐cell stage compared with that cultured in 6‐DMAP.     

Pharmacokinetics of intravenous and transdermal fentanyl in alpacas

The purpose of the study was to determine pharmacokinetics of fentanyl after intravenous (i.v.) and transdermal (t.d.) administration to six adult alpacas. Fentanyl was administered i.v. (2 μg/kg) or t.d. (nominal dose: 2 μg kg^?1 hr^?1). Plasma concentrations were determined using liquid chromatography–mass spectrometry. Heart rate and respiratory rate were assessed. Extrapolated, zero‐time plasma fentanyl concentrations were 6.0 ng/ml (1.7–14.6 ng/ml) after i.v. administration, total plasma clearance was 1.10 L hr^?1 kg^?1 (0.75–1.40 L hr^?1 kg^?1), volumes of distribution were 0.30 L/kg (0.10–0.99 L/kg), 1.10 L/kg (0.70–2.96 L/kg) and 1.5 L/kg (0.8–3.5 L/kg) for V₁, V₂, and V_ss, respectively. Elimination half‐life was 1.2 hr (0.5–4.3 hr). Mean residence time (range) after i.v. dosing was 1.30 hr (0.65–4.00 hr). After t.d. fentanyl administration, maximum plasma fentanyl concentration was 1.20 ng/ml (0.72–3.00 ng/ml), which occurred at 25 hr (8–48 hr) after patch placement. The area under the plasma fentanyl concentration‐vs‐time curve (extrapolated to infinity) after t.d. fentanyl was 61 ng*hr/ml (49–93 ng*hr/ml). The dose‐normalized bioavailability of fentanyl from t.d. fentanyl in alpacas was 35.5% (27–64%). Fentanyl absorption from the t.d. fentanyl patch into the central compartment occurred at a rate of approximately 50 μg/hr (29–81 μg/hr) between 8 and 72 hr after patch placement.     

Antimicrobial susceptibility testing of Spanish field isolates of Brachyspira hyodysenteriae

This study is the first conducted in Spain to evaluate antimicrobial susceptibility of field isolates of Brachyspira hyodysenteriae. One hundred and eight isolates of the bacterium, recovered from different Spanish swine farms between 2000 and 2007, were investigated. The minimum inhibitory concentrations (MIC) of erythromycin, tylosin, tiamulin, valnemulin, clindamycin and lincomycin were determined using a broth microdilution technique. Most of the isolates showed poor susceptibility to erythromycin (MIC₉₀ > 256 μg/ml), tylosin (MIC₉₀ > 256 μg/ml), clindamycin (MIC₉₀ > 4 μg/ml) and lincomycin (MIC₉₀ = 128 μg/ml). Reduced susceptibility to tiamulin and valnemulin was observed with a MIC > 2 μg/ml in 17.6% and 7.41% of the B. hyodysenteriae isolates, respectively. Moreover, a survival analysis permitted the detection of an increasing trend in the MIC values for almost all the antimicrobials used in the treatment of swine dysentery when comparing recent isolates (from 2006 to 2007) with those recovered in earlier years (between 2000 and 2004).     

Presence of nitric oxide synthase immunoreactivity and mRNA in buffalo (Bubalus bubalis) oocytes and embryos

This study examined the presence of immunoreactivity and mRNA for different nitric oxide synthase (NOS) isoforms in immature and in vitro matured oocytes and in embryos at two‐, four‐ and eight‐cell, and morula and blastocyst stages in buffalo. Oocytes obtained from slaughterhouse buffalo ovaries were subjected to in vitro maturation in TCM‐199 + 10% FBS + 5 μg/ml pFSH + 1 μg/ml estradiol‐17β + 0.81 mm sodium pyruvate + 10% buffalo follicular fluid + 50 μg/ml gentamycin sulphate for 24 h in a CO₂ incubator (5% CO₂ in air) at 38.5°C. Following in vitro fertilization carried out by incubating them with 2–4 million spermatozoa/ml for 18 h, the presumed zygotes were cultured in mCR2aa medium containing 0.6% BSA and 10% FBS for up to 8 days post insemination. Immunofluorescence staining of NOS using antibodies that cross‐reacted either with all the NOS isoforms i.e., universal (uNOS) or specifically with inducible (iNOS) or endothelial (eNOS) isoforms revealed that NOS was present in oocytes and embryos at all the stages examined. Examination of the semi‐quantitative expression of NOS genes by RT‐PCR revealed that the iNOS, eNOS and nNOS mRNA was present in the immature and mature oocytes and in all the embryonic stages examined. In conclusion, it was demonstrated in the present study that immunoreactivity and mRNA for different NOS isoforms was present in buffalo oocytes and pre‐implantation stage embryos.     

Immunomodulatory effect of Artemisia argyi polysaccharide on peripheral blood leucocyte of broiler chickens

This study was conducted to investigate the immunomodulatory effect of a water‐soluble polysaccharide extracted from Artemisia argyi (AAP) in vitro. The effect was assessed in peripheral blood leucocytes (PBLs) of broilers, which were incubated with different AAP concentrations (0, 25, 50, 100, and 200 μg/ml) for 24 hr at 37°C in a 5% CO₂ incubator. The results showed that, compared with the control group, immunoglobulin M (IgM) concentration was increased in the supernatant of the 100 μg/ml AAP‐treated group (p < .05), and immunoglobulin G (IgG) concentration was increased in the supernatant of the 200 μg/ml of AAP group (p < .05). In terms of cytokine production, production of interleukin‐1beta (IL‐1β), interleukin‐6 (IL‐6) and tumour necrosis factor‐alpha (TNF‐α) in the supernatant was enhanced in the AAP group in a dose‐dependent function, as well as enhanced mRNA expressions were showed in the cells (p < .05). The highest concentration of these three cytokines was observed in different AAP groups (IL‐1β for 25 μg/ml of AAP, IL‐6 for 100, and 200 μg/ml of AAP, and TNF‐α for 100 μg/ml of AAP respectively). The concentration of nitric oxide (NO) was increased when using AAP at the concentration of 100 μg/ml (p < .05) as compared to the control group. No significant effects on inducible nitric oxide synthase, Toll‐like receptor 4 (TLR4), myeloid differentiation factor 88 and nuclear factor Kappa B (NF‐κB) mRNA level were observed at each concentration of AAP. In conclusion, we found that AAP can specifically promote the production of immunoglobulins (IgM and IgG), cytokines (IL‐1β, IL‐6 and TNF‐α), as well as the NO concentration in vitro, but not through the activation of the TLR4/NF‐κB signalling pathway.     

Pharmacokinetics,disposition, and plasma concentrations of dimethyl sulfoxide (DMSO) in the horse following topical,oral, and intravenous administration

Compartmental models were used to investigate the pharmacokinetics of intravenous (i.v. ), oral (p.o. ), and topical (TOP ) administration of dimethyl sulfoxide (DMSO ). The plasma concentration–time curve following a 15‐min i.v. infusion of DMSO was described by a two‐compartment model. Median and range of alpha (t _1/2α) and beta (t _1/2β) half‐lives were 0.029 (0.026–0.093) and 14.1 (6.6–16.4) hr, respectively. Plasma concentration–time curves of DMSO following p.o. and TOP administration were best described by one‐compartment absorption and elimination models. Following the p.o. administration, median absorption (t _1/2ab) and elimination (t _1/2e) half‐lives were 0.15 (0.01–0.77) and 15.5 (8.5–25.2) hr, respectively. The plasma concentrations of DMSO were 47.4–129.9 μg/ml, occurring between 15 min and 4 hr. The fractional absorption (F ) during a 24‐hr period was 47.4 (22.7–98.1)%. Following TOP administrations, the median t _1/2ab and t _1/2e were 1.2 (0.49–2.3) and 4.5 (2.1–11.0) hr, respectively. Plasma concentrations were 1.2–8.2 μg/ml occurring at 2–4 hr. Fractional absorption following TOP administration was 0.48 (0.315–4.4)% of the dose administered. Clearance (Cl) of DMSO following the i.v. administration was 3.2 (2.2–6.7) ml hr^?1 kg^?1. The corrected clearances (Cl_F ) for p.o. and TOP administrations were 2.9 (1.1–5.5) and 4.5 (0.52–18.2) ml hr^?1 kg^?1.     

Hanging Drop Monoculture for Selection of Optimal Antioxidants During In Vitro Maturation of Porcine Oocytes

We analysed the effect of three antioxidants that have different functional mechanisms on the in vitro maturation (IVM) of porcine oocytes. Single oocyte monoculture using the hanging drop (HD) system has some advantages such as improving analysis efficiency brought by the smaller number of samples than the number of oocytes cultured in one drop. Direct effects of ligands on single oocytes could also be detected without considering the effects of paracrine factors from other oocytes. After 22 h of pre‐culture, denuded oocytes were cultured for 22 h with 0.01 and 0.1 μg/ml of L‐carnitine (LC), lactoferrin (LF) or sulforaphane (SF) in the presence/non‐presence of oxidant stress induced by H₂O₂ supplementation to evaluate the reducing effects against oxidative stress on nuclear maturation. As a result, compared with LC and SF, LF showed effective reduction in oxidative stress at a lower concentration (0.01 μg/ml), suggesting that LF is a more effective antioxidant in porcine oocyte IVM. Additionally, LF also increased maturation rate even in culture without H₂O₂. Our results clearly suggest that the HD monoculture system is useful for screening the substances that affect porcine oocyte culture.     

Does the Amount of Progesterone in Intravaginal Implants Used to Synchronise Oestrus Affect the Reproductive Performance of Brahman Heifers Artificially Inseminated at a Fixed Time

The study tested the hypothesis that reduced intravaginal implant progesterone (P₄) concentration to synchronise oestrus would increase pregnancy rates to fixed‐time artificial insemination (FTAI) in Bos indicus heifers. Brahman heifers (n = 294; 2 year) were body condition scored (BCS), weighed and scanned for presence of a corpus luteum (CL). Only cyclic heifers were selected and allocated randomly within BCS and 25 kg bodyweight category to one of three P₄ treatment groups. On day 10, heifers received a P₄ implant (CueMate‐1‐pod, 0.78g P₄; CueMate‐2‐pod, 1.56g P₄; or CIDR‐B, 1.9g P₄), 2 mg oestradiol benzoate (ODB) intramuscularly (IM) and 250 ug cloprostenol IM. At day 2, the implant was removed, 250 ug cloprostenol was injected IM and tail paint applied. The heifers received 1 mg ODB 24 h later and were FTAI 48–54 h after implant removal (day 0). Ten randomly selected heifers per group were blood sampled and scanned at days 10, 2, 0 and 6 to define the P₄ profiles pre‐ and post‐FTAI. Heifers were heat‐detected 18–20 days post‐FTAI and oestrous heifers AI’d by the AM/PM rule. Bulls joined the heifers on day 27 post‐FTAI. Transrectal ultrasonography estimated conception date on day 72. Statistical analysis examined the effects of treatment, technician, semen, ovarian status, BCS and liveweight, on pregnancy rate (PR) to FTAI. There was no significant difference (p = 0.362) in PR between treatment groups (CueMate 1‐pod, 36.4%; CueMate 2‐pod, 39.6%: CIDR‐B, 28.3%), but PR was higher in those heifers with increased BCS between FTAI and pregnancy diagnosis (p = 0.005). Thirty‐three per cent of monitor heifers had plasma P₄ concentrations of <1 ng/ml on day 6 after FTAI; only 20% of these conceived vs 60% of heifers with P₄ ≥ 1 ng/ml. In summary, no significant difference in PR was identified between treatments but good BCS and a rising plane of nutrition were critical to PR of these pure grade Brahman heifers in northern Australia.     

Effects of age and zinc supplementation on transport properties in the jejunum of piglets

Zinc is effective in the prevention and treatment of post‐weaning diarrhoea and in promoting piglet growth. Its effects on the absorption of nutrients and the secretory capacity of the intestinal epithelium are controversial. We investigated the effects of age, dietary pharmacological zinc supplementation and acute zinc exposure in vitro on small‐intestinal transport properties of weaned piglets. We further examined whether the effect of zinc on secretory responses depended on the pathway by which chloride secretion is activated. A total of 96 piglets were weaned at 26 days of age and allocated to diets containing three different levels of zinc oxide (50, 150 and 2500 ppm). At the age of 32, 39, 46 and 53 days, piglets were killed, and isolated epithelia from the mid‐jejunum were used for intestinal transport studies in conventional Ussing chambers, with 23 μm ZnSO₄ being added to the serosal side for testing acute effects. Absorptive transport was stimulated by mucosal addition of d ‐glucose or l ‐glutamine. Secretion was activated by serosal addition of prostaglandin E₂, carbachol or by mucosal application of Escherichia coli heat‐stable enterotoxin (St_p). Jejunal transport properties showed significant age‐dependent alterations (p < 0.03). Both absorptive and secretory responses were highest in the youngest piglets (32 d). The dietary zinc supplementation had no significant influence on jejunal absorptive and secretory responses. However, the pre‐treatment of epithelia with ZnSO₄ in vitro led to a small but significant decrease in both absorptive and secretory capacities (p < 0.05), with an exception for carbachol (p = 0.07). The results showed that, in piglets, chronic supplementation with zinc did not sustainably influence the jejunal transport properties in the post‐weaning phase. Because transport properties are influenced by the addition of zinc in vitro, we suggest that possible epithelial effects of zinc depend on the acute presence of this ion.