Carbapenem- and colistin-resistant Enterobacter has been a clinical and therapy problem in recent years. Here, we report the carbapenem- and colistin-resistant Enterobacter harboring blaIMI isolated from intestinal samples and the environment of a duck farm in China. Four blaIMI-positive Enterobacter isolates were resistant to carbapenem and colistin. Three blaIMI subtypes were detected in different molecular categories of Enterobacter. The detection of the various IMI producers highlights the diversity of carbapenemases in a duck farm. Whole-genome sequencing demonstrated the blaIMI genes were present in chromosomes or plasmids in these strains. The conjugation experiment demonstrated the ability of blaIMI-carrying plasmid to transmit horizontally. The molecular evolution characteristics were examined through comparative genetic analysis. The study demonstrated the presence of chromosomal and plasmid blaIMI and the blaIMI-carrying plasmid exhibits a horizontal transmission between Enterobacter and Escherichia coli C600. The similar genetic content was discovered between two blaIMI-16-positive Enterobacter asburiae. In addition, a blaIMI-16-carrying plasmid is an IncFII(Yp) plasmid, and a substantial amount of mobile genetic elements were identified around blaIMI-16. The IS-like elements and IncFII(Yp) plasmid are significant in the propagation of blaIMI. Our study provides evidence for the transmission of diverse blaIMI genes in China and supplies additional reference data for blaIMI-positive antimicrobial-resistant Enterobacter. Routine surveys of blaIMI-positive Enterobacter from animal-raising environments must be given more focus

Antimicrobial resistance has become a global problem that poses great threats to human health.  Antimicrobials are widely used in broiler chicken production and consequently affect their gut microbiota and resistome.  To better understand how continuous antimicrobial use in farm animals alters their microbial ecology, we used a metagenomic approach to investigate the effects of pulsed antimicrobial administration on the bacterial community, antibiotic resistance genes (ARGs) and ARG bacterial hosts in the feces of broiler chickens.  Chickens received three 5-day courses of individual or combined antimicrobials, including amoxicillin, chlortetracycline and florfenicol.  The florfenicol administration significantly increased the abundance of mcr-1 gene accompanied by floR gene, while amoxicillin significantly increased the abundance of genes encoding the AcrAB-tolC multidrug efflux pump (marA, soxS, sdiA, rob, evgS and phoP).  These three antimicrobials all led to an increase in Proteobacteria.  The increase in ARG host, Escherichia, was mainly attributed to the β-lactam, chloramphenicol and tetracycline resistance genes harbored by Escherichia under the pulsed antimicrobial treatments.  These results indicated that pulsed antimicrobial administration with amoxicillin, chlortetracycline, florfenicol or their combinations significantly increased the abundance of Proteobacteria and enhanced the abundance of particular ARGs.  The ARG types were occupied by the multidrug resistance genes and had significant correlations with the total ARGs in the antimicrobial-treated groups.  The results of this study provide comprehensive insight into pulsed antimicrobial-mediated alteration of chicken fecal microbiota and resistome.

The objective of this study was to verify the supposition that efflux might be involved in the drug resistance of Riemerella anatipestifer isolates. Two broad-spectrum efflux pump inhibitors, carbonyl cyanide 3-chlorophenylhydrazone (CCCP) and Phe-Arg-β-naphthylamide (PAβN), on the contribution of minimum inhibitory concentrations of amikacin, streptomycin, chloramphenicol, tetracycline, ceftriaxone, ceftazidime, nalidixic acid, levofloxacin, enrofloxacin, as well as ciprofloxacin against 69 clinical R. anatipestifer isolates were investigated. We first reported that the two efflux pump inhibitors could restore the antimicrobial susceptibility of R. anatipestifer isolates. It is suggested that active efflux system is possible to be linked with the development of resistance in R. anatipestifer isolates.

Contamination of deoxynivalenol (DON) in grains is common worldwide and pigs are particularly susceptible to this mycotoxin. The distribution of DON in porcine tissues following intravenous administration was investigated in this study. Fifteen pigs were randomly divided into three groups. Animals in groups A and B were administrated with DON at the dose of 250 and 750 μg kg–1 body weight, respectively, while group C served as blank control. Plasma, bile and 27 tissues were collected at 30 min post-administration. DON concentrations in all samples were tested using high-performance liquid chromatography- tandem mass spectrometry (HPLC-MS/MS). To observe the distribution of DON in tissues, these samples were further subjected to the immunohistochemical analyses. Totally, the bile and 13 tissues were sampled for DON-based detection, including kidney, mesenteric lymph nodes, muscle, stomach, jejunum, colon, plasma, spleen, rectum, cecum, liver, ileum, and duodenum. No significant difference was observed for the concentrations of DON in duodenum, ileum and liver samples between groups A and B; while the DON concentrations in cecum and rectum of group B were significantly higher (P-value <0.05) than those in group A. In addition, the DON concentrations in stomach, jejunum, colon, mesenteric lymph nodes, muscle, kidney, spleen, bile, and plasma of group B were remarkably higher than those of group A (P-value<0.01). Levels of DON in other 14 tissues including medulla oblongata, midbrain, diencephalon, pons, tip and tongue body, tongue, soft palate, tonsils, pharyngeal mucosa, oral buccal mucosa, thymus, thyroid, esophagus and adrenal gland were all below the limit of detection. The results of immunohistochemistry showed that 11 tissue samples (medullaoblongata, tonsil, adrenal medulla, thyroid gland, thyroid, stomach, duodenum, jejunum, kidney, spleen, and mesenteric lymph nodes) were positive and DON was mainly distributed around blood vessels in these tissues. Therefore, we believed that concentrations of DON in tissues differ when pigs are in exposure to various dosages and DON causes lesions in many pig tissues.

The purpose of the study was to investigate the pharmacokinetics of cefquinome in plasma and milk samples of lactating Chinese Holstein following a single intramammary administration into one quarter at the dose of 75 mg. Residue depletion of cefquinome in milk administrated at one quarter following three consecutive infusions at the same dose were also carried out. Cefquinome concentrations in plasma and milk were determined by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method. A non-compartmental analysis was used to obtain the pharmacokinetic parameters of cefquinome. Following the single treatment, cefquinome wasn’t detected in any of the plasma samples. The concentration of cefquinome in milk reached peaked values (Cmax) of (599.00±322.00) μg mL-1 at 2 h after administration (Tmax), elimination half-life (t1/2λz) was (4.63±0.26) h, area under the concentration-time curve (AUC0-∞) was (4 890.19±1 906.98) μg mL-1 h, and mean residence time (MRT) was (6.03±2.27) h. In residue depletion study, cefquinome concentrations in 5 out of 6 milk samples at 72 h were lower than the maximum residue limit fixed by the European regulatory agency (20 μg kg-1 for cefquinome) and cefquinome still could be detected in milk of treated quarters at 120 h post-treatment. The maximum concentration (Cmax) of cefquinome in milk from treated quarters was (486.50±262.92) μg mL-1 and arrived at 6 h after administration (Tmax), elimination half-life (t1/2λz) was (6.30±0.76) h, area under the concentration-time curve (AUC0-∞) was (44747.79±11434.43) μg mL-1 h, and mean residence time (MRT) was (10.09±1.40) h. This study showed that cefquinome has the feature of poor penetration into blood and was eliminated quickly from milk in lactating cows after intramammary administration.

Pharmacokinetics of cyadox (CYX) and its major metabolites in healthy swine was investigated in this paper. 1,4- Bisdesoxycyadox (BDCYX), cyadox-1-monoxide (CYX-1-O) and quinoxaline-2-carboxylic acid (QCA), three main metabolites of cyadox, were synthesized by College of Science, China Agricultural University. Cyadox (CYX) was administered to 8 healthy cross-bread swine intravenously (i.v.) and orally (p.o.) at a dosage of 1 mg kg-1 body weight and 40 mg kg-1 body weight respectively in a randomized crossover design test with 2-wk washout period. A sensitive high-performance liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) method was developed for the determination of cyadox and its major metabolites in plasma. CYX and its major metabolites BDCYX, and CYX-1-O can be detected after intravenous administration of cyadox while CYX and its metabolites BDCYX, CYX-1-O and QCA can be detected after oral administration of CYX. Plasma concentration vs. time profiles of CYX and its major metabolites were analyzed by non-compartmental pharmacokinetic method. Following i.v. administration, the areas under the plasma concentration-time curve (AUC0- ) were (0.38±0.03) μg mL-1 h (CYX), (0.018±0.002) μg mL-1 h (BDCYX) and (0.17±0.02) μg mL-1 h (CYX-1-O), respectively. The terminal elimination half-lives (t1/2lz) were determined to be (0.93±0.07) h (CYX), (1.45±0.04) h (BDCYX), and (0.92±0.04) h (CYX-1-O), respectively. Steady-state distribution volume (Vss) of (2.14±0.11) L kg-1 and total body clearance (CL) of (2.84±0.19) L h-1 kg-1 were determined for CYX after i.v. dosing. The bioavailability (F) of CYX was 2.85% for oral administration. After single i.v. administration, peak plasma concentrations (Cmax) of (1.08±0.06) μg mL-1 (CYX), (0.0068± 0.0004) μg mL-1 (BDCYX) and (0.25±0.03) μg mL-1 (CYX-1-O) were observed at Tmax of 0.033 h (CYX), 1 h (BDCYX) and 0.033 h (CYX-1-O), respectively. The main pharmacokinetic parameters after p.o. administration were as follows: AUC0- were (0.42±0.04) μg mL-1 h (CYX), (1.38±0.14) μg mL-1 h (BDCYX), (0.59±0.02) μg mL-1 h (CYX-1-O) and (1.48±0.09) μg mL-1 h (QCA), respectively. t1/2lz were (4.77±0.33) h (CYX), (5.77±0.56) h (BDCYX), (4.12±0.28) h (CYX-1-O), and (8.51±0.39) h (QCA), respectively. After p.o. administration, Cmaxs of (0.033±0.002) μg mL-1 (CYX), (0.22±0.03) μg mL-1 (BDCYX), (0.089±0.005) μg mL-1 (CYX-1-O), and (0.17± 0.01) μg mL-1 (QCA) were observed at Tmax of (7.38±0.33) h (CYX), (7.25±0.31) h (BDCYX), (7.38±0.33) h (CYX-1-O), and (7.25±0.31) h (QCA), respectively. The results showed that CYX was slowly absorbed after oral administration and most of CYX was transformed to its metabolites in swine. The area under plasma concentration-time curve (AUC0- )of metabolites were higher than that of CYX after p.o. administration, and the elimination half-lives (t1/2lz) of QCA were longer than those of CYX, CYX-1-O, and BDCYX after oral administration.

An accurate and precise method for the determination of tulathromycin in swine plasma was developed and validated. Plasma samples were analyzed by high-performance liquid chromatography with tandem mass spectrometry detection (HPLC-MS/MS) using electrospray ionization (ESI). Tulathromycin was extracted from plasma by precipitation with acetonitrile and separated using a Phenomenex Luna 5 μm C18 column (150 mm×2.0 mm) at a flow rate of 0.25 mL min-1. Solvent A consisted of 0.002 mol L-1 ammonium acetate and formic acid (999:1, v/v), and solvent B was acetonitrile. The mass spectrometer was operated in the selected-ion mode with atmospheric pressure chemical ionization to monitor the respective MH+ ions, namely, m/z 577.3 for tulathromycin and m/z 679.3 for the internal standard roxithromycin. The calibration curves were linear in a dynamic range of 2.0-500 ng mL-1 on the column. The accuracy was ranged from 95.25 to 109.75%, and the precision was ranged from 2.81 to 7.72%. The recoveries measured at 3 concentration levels (20, 250, and 500 ng mL-1) were higher than 98%. The method described above is efficient, and has the required accuracy and precision for rapid determination of tulathromycin in plasma. The method was applied to study the pharmacokinetics of tulathromycin in swine, and tulathromycin demonstrated a rapid absorption, wide distribution, and slow elimination after intramuscular administration.