摘要

       我们之前的研究表明，特定的抗生素抗性基因 (ARG) 在芬兰北部波罗的海养鱼场下方的沉积物中局部富集，即使抗生素的选择压力可以忽略不计。我们假设养殖鱼粪的不断涌入可能是养殖场沉积物中富含 ARG 的合理来源。在本研究中，我们分析了来自波罗的海养殖场的 20 条鱼的肠道内容物的抗生素抗性组的组成。我们使用了一种高通量方法，即具有 364 个引物组的 WaferGen qPCR 阵列来检测和量化 ARG、移动遗传元件 (MGE) 和 16S rRNA 基因。尽管 qPCR 引物组的选择范围相当广泛，但在肠内容物中仅检测到 28 个基因。检测到的基因是编码对磺胺(sul1)、甲氧苄氨嘧啶(dfrA1)、四环素[tet(32)、tetM、tetO、tetW]、氨基糖苷(aadA1、aadA2)、氯霉素(catA1)和外排泵抗性基因的抗性基因ARGs （emrB、matA、mefA、msrA）。检测到的基因还包括1类整合子相关基因（intI1、qacEΔ1）和转座酶（tnpA）。重要的是，大多数检测到的基因与农场沉积物中富集的基因相同。这项初步研究表明，尽管养殖场缺乏同期抗生素治疗，但养殖鱼类的粪便有助于养殖场沉积物中的 ARG 富集。我们观察到个体养殖鱼类的肠道内容物有其自身的抵抗组成分。我们的结果还表明转座酶和 tet 基因的总相对丰度显着相关（p = 0.001，R2 = 0.71）。此外，我们分析了养殖鱼的粘膜皮肤和鳃丝抵抗组，但仅检测到一种多药外排抗性基因（emrB）。据我们所知，这是第一次使用独立于培养的方法报告养殖鱼类的抵抗力的研究。确定 ARG 的可能来源，尤其是动员的 ARG，对于控制 ARG 在养鱼设施中的发生和传播以及降低 ARG 从养殖场传播到周围环境的风险至关重要。

       Our previous studies showed that particular antibiotic resistance genes (ARGs) were enriched locally in sediments below fish farms in the Northern Baltic Sea, Finland, even when the selection pressure from antibiotics was negligible. We assumed that a constant influx of farmed fish feces could be the plausible source of the ARGs enriched in the farm sediments. In the present study, we analyzed the composition of the antibiotic resistome from the intestinal contents of 20 fish from the Baltic Sea farms. We used a high-throughput method, WaferGen qPCR array with 364 primer sets to detect and quantify ARGs, mobile genetic elements (MGE), and the 16S rRNA gene. Despite a considerably wide selection of qPCR primer sets, only 28 genes were detected in the intestinal contents. The detected genes were ARGs encoding resistance to sulfonamide (sul1), trimethoprim (dfrA1), tetracycline [tet(32), tetM, tetO, tetW], aminoglycoside (aadA1, aadA2), chloramphenicol (catA1), and efflux-pumps resistance genes (emrB, matA, mefA, msrA). The detected genes also included class 1 integron-associated genes (intI1, qacEΔ1) and transposases (tnpA). Importantly, most of the detected genes were the same genes enriched in the farm sediments. This preliminary study suggests that feces from farmed fish contribute to the ARG enrichment in farm sediments despite the lack of contemporaneous antibiotic treatments at the farms. We observed that the intestinal contents of individual farmed fish had their own resistome compositions. Our result also showed that the total relative abundances of transposases and tet genes were significantly correlated (p = 0.001, R2 = 0.71). In addition, we analyzed the mucosal skin and gill filament resistomes of the farmed fish but only one multidrug-efflux resistance gene (emrB) was detected. To our knowledge, this is the first study reporting the resistome of farmed fish using a culture-independent method. Determining the possible sources of ARGs, especially mobilized ARGs, is essential for controlling the occurrence and spread of ARGs at fish farming facilities and for lowering the risk of ARG spread from the farms to surrounding environments.

https://www.frontiersin.org/articles/10.3389/fmicb.2016.02137/full