Institute of intelligent machinery, Hefei Institute of Material Sciences, Chinese Academy of Sciences, Hefei 230031, China.
Department of Biosystems Engineering, University of Manitoba, Winnipeg, MB R3T 5V6, Canada.
Int J Environ Res Public Health. 2020 Mar 17;17(6):1973. doi: 10.3390/ijerph17061973.
(1) Objective: The objective of this study was to screen amoxicillin (AMX)-degrading bacterial strains in pig manure and optimize the fermentation conditions for these strains to achieve high fermentation rate, which can provide an effective way for the practical application of bacterial strains as antibiotic-degrading bacterial in treating livestock waste for antibiotic residues. (2) Methods: Antibiotic susceptibility tests and high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) were employed to screen AMX-degrading bacterial strains in pig manure. The culture conditions were optimized for AMX-degrading bacterial strains using Plackeet-Burman design (PBD), the steepest ascent design, and the response surface methods, coupled with the Box-Behnken design (BBD). The effects of culture time, temperature, rotator (mixing) speed, inoculum level, and initial pH value on the growth of AMX-degrading strains were investigated. Experimental data obtained from BBD were utilized to generate a second-order polynomial regression model for evaluating the effects of the tested variables on the optical density at 600 nm (OD) of culture solutions as the growth indicator for the screened AMX-degrading strains. (3) Results: The initial pH, culture time, and the inoculum level had significant effects on the OD value (growth) of the screened AMX-degrading strains. The initial pH value was found to be the most critical factor influencing the growth of bacteria. The optimized culture condition for the bacterial growth determined by the response surface methodology was: the initial pH of 6.9, culture time of 52 h, and inoculum level of 2%. The average OD value of 12 different fermentation conditions in the initial fermentation tests in this study was 1.72 and the optimization resulted in an OD value of 3.00. The verification experiment resulted in an OD value of 2.94, which confirmed the adequacy of the optimization model for the determining the optimal culture condition. (4) Conclusions: The growth of the screened strain of AMX-degrading bacteria could be optimized by changing the fermentation conditions. The optimization could be achieved by using the Box-Behnken response surface method and Plackett-Burman experimental design.
(1) 目的：本研究旨在筛选猪粪中降解阿莫西林（AMX）的细菌菌株，并优化这些菌株的发酵条件，以实现高发酵率，为将细菌菌株作为抗生素降解菌实际应用于处理含有抗生素残留的牲畜废物提供有效途径。(2) 方法：采用抗生素药敏试验和高效液相色谱串联质谱（HPLC-MS/MS）筛选猪粪中 AMX 降解细菌菌株。采用 Plackett-Burman 设计（PBD）、最陡爬坡设计和响应面法对 AMX 降解细菌菌株的培养条件进行优化，并结合 Box-Behnken 设计（BBD）对培养时间、温度、搅拌器（混合）速度、接种量和初始 pH 值对 AMX 降解菌生长的影响进行研究。利用 BBD 获得的实验数据，建立二次多项式回归模型，评估测试变量对筛选出的 AMX 降解菌培养溶液在 600nm 处的光密度（OD）的影响，作为筛选出的 AMX 降解菌生长的指标。(3) 结果：初始 pH 值、培养时间和接种量对筛选出的 AMX 降解菌的 OD 值（生长）有显著影响。结果发现，初始 pH 值是影响细菌生长的最关键因素。响应面法确定的细菌生长最佳培养条件为：初始 pH 值 6.9、培养时间 52h、接种量 2%。本研究初始发酵试验中 12 种不同发酵条件的平均 OD 值为 1.72，优化后 OD 值为 3.00。验证试验的 OD 值为 2.94，表明优化模型确定最佳培养条件是合理的。(4) 结论：通过改变发酵条件可以优化筛选出的 AMX 降解菌的生长。可以采用 Box-Behnken 响应面法和 Plackett-Burman 实验设计来实现优化。
