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The co-exposure of antibiotics has important effects on antibiotic resistance genes (ARGs) and microbial community aggregation in wastewater treatment plants (WWTPs). However, it is unclear whether differences in historical antibiotic exposure stress can determine responses of microbes and ARGs to combined antibiotics. By selecting a high concentration (30 mg·L -1 ) of sulfamethoxazole (SMX) and trimethoprim (TMP) as historical exposure stress conditions, the effects of SMX and TMP-combined pollution on ARGs, bacterial communities, and their interactions were explored in short-term experiments. Based on high-throughput quantitative PCR, a total of 13 ARGs were detected, and the absolute abundance was 2.21-5.42 copies·μL -1 (logarithm, DNA, the same below). Among them, sul2, ermB, mefA , and tetM-01 were the main subtypes in the samples, and the absolute abundance was between 2.95 and 5.40 copies·μL -1 . The combined contamination of SMX and TMP could cause the enrichment of ARGs and mobile genetic elements (MGEs); however, their effects on each subtype were different, and the historical legacy effect of SMX was higher than that of TMP. Under the different exposure histories, the co-occurrence and co-exclusion patterns existed between ARGs. Moreover, MGEs (especially intI-1 ) were significantly correlated with sulfonamides ( sul1 and sul2 ), tetracyclines[ tet(32) ], and macrolide-lincosamide-streptogramin (MLSB) resistance genes ( ermB ). Based on the full-scale classification of microorganisms, it was found that the microbial community structure of various groups responded differently to combined pollution, and the conditionally abundant taxa (CAT) were obviously enriched. Thauera, Pseudoxanthomonas, and Paracoccus were the dominant resistant bacterial genera. Furthermore, a total of 31 potential hosts of ARGs were identified with network analysis, which were dominated with conditionally rare taxa (CRT). Particularly, Candidatus_Alysiosphaera and Fusibacter were positively correlated with most of the ARGs, being the common protentional hosts. Importantly, some rare genera (RT, Variibacter, Aeromonas, Cloacibacterium , etc.) were potential hosts of transposon IS613 , which played an important role in the proliferation and spread of ARGs. In conclusion, this study revealed the legacy effects of historical antibiotic stress on ARGs and their hosts, which could provide new ideas and theoretical basis for reducing ARGs pollution in WWTPs.

To study the enzyme kinetics of ligustilide metabolism and the effects of selective CYP450 inhibitors on the metabolism of ligustilide in liver microsomes of rat, a LC-MS method was established for quantitative analysis of ligustilide in liver microsomes incubation system with nitrendipine as internal standard. The determination m/z for ligustilide was 173, and for nitrendipine, 315. An optimum incubation system was found and various selective CYP inhibitors were used to investigate their inhibitory effects on the metabolism of ligustilide. The results showed that enzyme kinetics of ligustilide could be significantly inhibited by ketoconazole, trimethoprim and a-naphthoflavon but scarcely inhibited by omeprazole, 4-methylpyrazole and quinidine. Therefore, CYP3A4, CYP2C9 and CYP1A2 are the major isoenzyme participated in in vitro metabolism of ligustilide.

Objective: To investigate the antibiotics-resistance type and molecular epidemiology of Streptococcus pneumoniae isolated from children in Hangzhou., Methods: The sensitivities of 323 strains of Streptococcus pneumoniae to 9 antibiotics were determined in vitro by Kirby-Bauer diffuse methods, and MICs of penicillin and cefotaxime were determined by E-test methods., Results: Among all 323 strains isolated from children during the period from August 2001 to July 2002, 136 strains (42.1%) were sensitive to penicillin, while 57 strains (17.7%) were penicillin-resistant. Penicillin MICs ranged from 0.012 microg/ml to 4.0 microg/ml. All the strains were sensitive to cefotaxime and its MICs ranged from 0.012 microg/ml to 4.0 microg/ml. The most resistant antibiotic was erythromycin and it's resistant-rate was as high as 90.7%, followed by tetracycline (87.6%), trimethoprim-sulfamethoxazole (48.6%) and chloromycetin (14.9%). Totally 197 strains (61.0%) were multi-drug-resistant pneumococci and most of them were resistant to trimethoprim-sulfamethoxazole, erythromycin and tetracycline at the same time. Two strains (0.6%) were resistant to rifampin and none was resistant to vancomycin and ofloxacin. BOX PCR typing was carried out and no overwhelming fingerprinting pattern was found among penicillin resistant Streptococcus pneumoniae strains which were isolated from patients, while the banding patterns were always similar or identical among the strains isolated from the same specimen or from the same patient at different time, respectively., Conclusion: The antibiotics-resistant rate of pneumococci was high in Hangzhou, but the third-generation cephalosporins were still the best antibiotics against Streptococcus pneumoniae. One child could be infected or colonized by more than one pneumococci clone at the same or different time.

The agar-dilution method was adopted to study contrastively the bacteriostasis of Rhizoma Coptidis and TMP, and also of the two drugs in combination. The results indicate that the combination works in good cooperation against Escherichia coli, Bacillus pyocyaneus, Staphylococcus aureus, Shigella flexneri and S. dysenteriae.