Search

To evaluate the potential effects of antibiotics on ammonia-oxidizing microbes, multiple tools including quantitative PCR (qPCR), 454-pyrosequencing, and a high-throughput functional gene array (GeoChip) were used to reveal the distribution of ammonia-oxidizing archaea (AOA) and archaeal amoA (Arch-amoA) genes in three wastewater treatment systems receiving spiramycin or oxytetracycline production wastewaters. The qPCR results revealed that the copy number ratios of Arch-amoA to ammonia-oxidizing bacteria (AOB) amoA genes were the highest in the spiramycin full-scale (5.30) and pilot-scale systems (1.49 × 10(-1)), followed by the oxytetracycline system (4.90 × 10(-4)), with no Arch-amoA genes detected in the control systems treating sewage or inosine production wastewater. The pyrosequencing result showed that the relative abundance of AOA affiliated with Thaumarchaeota accounted for 78.5-99.6% of total archaea in the two spiramycin systems, which was in accordance with the qPCR results. Mantel test based on GeoChip data showed that Arch-amoA gene signal intensity correlated with the presence of spiramycin (P < 0.05). Antibiotics explained 25.8% of variations in amoA functional gene structures by variance partitioning analysis. This study revealed the selection of AOA in the presence of high concentrations of spiramycin in activated sludge systems.

To monitor spiramycin (SP) residue in milk, a monoclonal antibody (mAb)-based indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed. This study described the preparation of three immunogens and the production of a high-affinity mAb. After optimization, the 50% inhibition concentration (IC50) for the developed icELISA was estimated as 0.97 ng/mL in the assay buffer, and the limit of detection and limit of quantitation were 2.51 and 4.40 μg/L in the milk matrix. The newly developed assay demonstrated negligible cross-reactivity with 15 other macrolide antibiotics, but not with kitasamycin (23.4%). The mean recoveries ranged from 81 to 103% for the spiked samples (5, 10, and 50 μg/L), and the coefficient of variation ranged from 5.4 to 9.6%. The icELISA was validated by LC-MS/MS method, and all results demonstrated that it was a suitable screening method for detecting SP residue in milk without requiring a cleanup process.

A method using liquid chromatography electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) for the determination of trace levels of five macrolide antibiotics (spiramycin, tilmicosin, oleandomycin, erythromycin, and tylosin) in eggs is presented. Data acquisition under MS/MS was achieved by applying multiple reaction monitoring (MRM) of two or three fragment ion transitions to provide a high degree of sensitivity and specificity for both quantification and confirmation. Matrix-matched standard calibration curves were used to achieve the best accuracy of the method. A fully nested experimental design was used to study the measurement uncertainty arising from intermediate precision and trueness or proportional bias. The overall recoveries, that is, those determined by the nested experiments, of spiramycin, tilmicosin, oleandomycin, erythromycin, and tylosin at fortified levels of 60, 100, 200, and 300 microg/kg were 96.8, 98.2, 98.3, 98.8, and 95.4%, respectively. The LC/ESI-MS/MS method detection limits (S/N > or = 3:1) of five macrolides were <1.0 microg/kg.

Liquid chromatography-electrospray ionization mass spectrometry methods (LC-ESI-MS and LC-ESI-MS/MS) for the determination of five macrolide antibiotics including spiramycin, tilmicosin, oleandomycin, erythromycin, and tylosin in honey are presented. Macrolides were protonated to form singly and/or doubly charged pseudomolecular ions, depending on their chemical structures, in an electrospray positive ionization mode. Data acquisition under MS/MS was achieved by applying multiple reaction monitoring (MRM) of two or three fragment ion transitions to provide a high degree of sensitivity and specificity. The recoveries, that is, determined by LC-ESI-MS/MS, of the five macrolides at fortified levels of 6, 16, 40, and 80 microg/kg ranged from 75.5 to 135.7% in light honey and from 42.1 to 111.0% in dark honey. The ion ratios obtained under MS/MS were key criteria to confirm the identity of macrolides in incurred samples. LC-ESI-MS/MS method detection limits of the five macrolides were <0.1 microg/kg.