Université de Lorraine, Laboratoire Interdisciplinaire des Environnements Continentaux, UMR 7360, 54506 Vandoeuvre-lès-Nancy, France.
Res Microbiol. 2013 Jun;164(5):457-65. doi: 10.1016/j.resmic.2013.01.012. Epub 2013 Feb 6.
We describe a real-time PCR assay for the quantitative detection of arsB and ACR3(1) arsenite transporter gene families, two ubiquitous and key determinants of arsenic resistance in prokaryotes. The assay was applied in batch growth experiments using a wasteland soil bacterial community as an inoculum to investigate the effect of increasing arsenite [As(III)] concentrations on genes and transcript abundances. The aioA gene encoding the large subunit of arsenite oxidase was monitored in parallel. Results showed that arsB and ACR3(1) gene abundances correlated positively with the As(III) concentration. Both genes showed similar transcription patterns and strong upregulation by arsenic. Microbial As(III) oxidation occurred in As(III) spiked cultures and was associated with expression of the aioA gene in most cases. However, aioA was also expressed in several non-amended culture replicates. Analysis of cDNA clone libraries revealed that Pseudomonas was the dominant metabolically active genus whatever the As(III) concentration. Expressed arsB and ACR3(1) gene sequences were also affiliated with those from Pseudomonas, while expressed aioA sequences were more taxonomically diverse. The study suggests that arsenite transporter genes are appropriate biomarkers of arsenic stress that may be suitable for further exploring the adaptive response of bacterial communities to arsenic in contaminated environments.
我们描述了一种实时 PCR 检测方法,用于定量检测 ArsB 和 ACR3(1)砷酸盐转运基因家族,这两个基因家族是原核生物中砷抗性的普遍且关键决定因素。该检测方法应用于批生长实验中,使用荒地土壤细菌群落作为接种物,以研究增加亚砷酸盐 [As(III)] 浓度对基因和转录物丰度的影响。同时监测编码亚砷酸盐氧化酶大亚基的 aioA 基因。结果表明,arsB 和 ACR3(1)基因丰度与 As(III)浓度呈正相关。这两个基因表现出相似的转录模式,并且受到砷的强烈上调。在添加亚砷酸盐的培养物中发生了微生物亚砷酸盐氧化作用,并且在大多数情况下与 aioA 基因的表达相关。然而,在几个未经修饰的培养物重复中也表达了 aioA。cDNA 克隆文库分析表明,无论 As(III)浓度如何,假单胞菌都是主要的代谢活跃属。表达的 arsB 和 ACR3(1)基因序列也与假单胞菌的序列相关,而表达的 aioA 序列则具有更多的分类多样性。该研究表明,砷酸盐转运基因是砷胁迫的合适生物标志物,可能适合进一步探索细菌群落对污染环境中砷的适应性反应。