Li Qing, Chang Weishan, Zhang Hongna, Hu Dong, Wang Xuepeng
Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Shandong Agricultural University, Tai'an, China.
Laboratory for Marine Fisheries Science and Food Production Processes, Qingdao National Laboratory for Marine Science and Technology, Qingdao, China.
Front Microbiol. 2019 Apr 3;10:633. doi: 10.3389/fmicb.2019.00633. eCollection 2019.
We compared the diversity of extended-spectrum β-lactamases (ESBLs) producing () in wastewater of a municipal wastewater treatment plant. This was done by analyzing multiple antibiotic resistant phenotypes and genotypes. Also, we investigated the antibiotic resistance transfer mechanism of the plasmid by comparing the antibiotic resistance gene linked transfer using a conjugative test, and by analyzing the full-length DNA sequence of one plasmid. The results showed that 50 ESBLs-producing isolates were isolated from 80 wastewater samples at the rate of 62.5% (50/80), out of which 35 transconjugants were obtained with the multiple antibiotic resistant transfer rate as high as 70.0% (35/50). Multiple antibiotic resistance was shown in all transconjugants and donor bacteria, which were capable of resistance to 11 out of 15 kinds of antibiotics. Both transconjugants and donors were capable of resistance to the Ampicillin and Cefalotin at a rate of 100.00% (35/35), while the total antibiotic resistant spectrum of transconjugants narrowed at the rate of 94.29% (33/35) and broadened at the rate of 5.71% (2/35) after conjugate to the donor bacteria. PCR showed that the resistant genotypes decreased or remained unchanged when compared to donor bacteria with transconjugants while the and genes were transferred and linked at a rate of 100.00% (35/35) and the gene was at the rate as high as 94.29% (33/35). However, the gene was transferred at a low rate of 4.17% (1/24). In addition, the major resistance gene subtypes were , , and according to sequencing and Blast comparison. Plasmid wwA8 is a closed-loop DNA molecule with 83157 bp, and contains 45 predicted genes, including three antibiotic resistant resistance genes, , and , which can be transferred with . This study shows that isolated from wastewater was capable of transferring resistance genes and producing antibiotic resistant phenotypes. The plasmids containing different resistance genes in play an important role in the multiple antibiotic resistant transfer. Most importantly, antibiotic resistant resistance genes have different transfer efficiencies, the and genes transferred at a rate of 100.00% and linked transfer in all 35 transconjugants.
我们比较了城市污水处理厂废水中产超广谱β-内酰胺酶（ESBLs）的（）的多样性。通过分析多种抗生素耐药表型和基因型来完成此项工作。此外，我们通过接合试验比较抗生素耐药基因连锁转移，并分析一个质粒的全长DNA序列，研究了质粒的抗生素耐药转移机制。结果显示，从80份废水样本中分离出50株产ESBLs的（），分离率为62.5%（50/80），其中获得35株接合子，多重抗生素耐药转移率高达70.0%（35/50）。所有接合子和供体细菌均表现出多重抗生素耐药，能够对15种抗生素中的11种产生耐药。接合子和供体对氨苄西林和头孢噻吩的耐药率均为100.00%（35/35），而与供体细菌接合后，接合子的总抗生素耐药谱以94.29%（33/35）的比例变窄，以5.71%（2/35）的比例变宽。PCR结果显示，与供体细菌相比，接合子的耐药基因型减少或保持不变，而（）和（）基因的转移和连锁率为100.00%（35/35），（）基因的转移率高达94.29%（33/35）。然而，（）基因的转移率较低，为4.17%（1/24）。此外，根据测序和Blast比较，主要的耐药基因亚型为（）、（）和（）。质粒wwA8是一个83157 bp的闭环DNA分子，包含45个预测基因，其中包括三个抗生素耐药基因，（）、（）和（），它们可与（）一起转移。本研究表明，从废水中分离出的（）能够转移耐药基因并产生抗生素耐药表型。（）中含有不同耐药基因的质粒在多重抗生素耐药转移中起重要作用。最重要的是，抗生素耐药基因具有不同的转移效率，（）和（）基因的转移率为100.00%，并在所有35株接合子中连锁转移。
