Card Roderick M, Warburton Philip J, MacLaren Nikki, Mullany Peter, Allan Elaine, Anjum Muna F
Department of Bacteriology, Animal Health and Veterinary Laboratories Agency, Addlestone, Surrey, United Kingdom.
Department of Microbial Diseases, Eastman Dental Institute, University College London, London, United Kingdom.
PLoS One. 2014 Jan 22;9(1):e86428. doi: 10.1371/journal.pone.0086428. eCollection 2014.
The aim of this study was to screen for the presence of antimicrobial resistance genes within the saliva and faecal microbiomes of healthy adult human volunteers from five European countries. Two non-culture based approaches were employed to obviate potential bias associated with difficult to culture members of the microbiota. In a gene target-based approach, a microarray was employed to screen for the presence of over 70 clinically important resistance genes in the saliva and faecal microbiomes. A total of 14 different resistance genes were detected encoding resistances to six antibiotic classes (aminoglycosides, β-lactams, macrolides, sulphonamides, tetracyclines and trimethoprim). The most commonly detected genes were erm(B), blaTEM, and sul2. In a functional-based approach, DNA prepared from pooled saliva samples was cloned into Escherichia coli and screened for expression of resistance to ampicillin or sulphonamide, two of the most common resistances found by array. The functional ampicillin resistance screen recovered genes encoding components of a predicted AcrRAB efflux pump. In the functional sulphonamide resistance screen, folP genes were recovered encoding mutant dihydropteroate synthase, the target of sulphonamide action. The genes recovered from the functional screens were from the chromosomes of commensal species that are opportunistically pathogenic and capable of exchanging DNA with related pathogenic species. Genes identified by microarray were not recovered in the activity-based screen, indicating that these two methods can be complementary in facilitating the identification of a range of resistance mechanisms present within the human microbiome. It also provides further evidence of the diverse reservoir of resistance mechanisms present in bacterial populations in the human gut and saliva. In future the methods described in this study can be used to monitor changes in the resistome in response to antibiotic therapy.
本研究的目的是筛查来自五个欧洲国家的健康成年人类志愿者唾液和粪便微生物群中抗菌抗性基因的存在情况。采用了两种非培养方法来避免与微生物群中难以培养的成员相关的潜在偏差。在基于基因靶点的方法中，使用微阵列来筛查唾液和粪便微生物群中70多种临床上重要的抗性基因的存在情况。总共检测到14种不同的抗性基因，它们编码对六种抗生素类别（氨基糖苷类、β-内酰胺类、大环内酯类、磺胺类、四环素类和甲氧苄啶）的抗性。最常检测到的基因是erm(B)、blaTEM和sul2。在基于功能的方法中，将从混合唾液样本中制备的DNA克隆到大肠杆菌中，并筛选对氨苄青霉素或磺胺类药物的抗性表达，这是阵列检测到的两种最常见的抗性。功能性氨苄青霉素抗性筛选回收了编码预测的AcrRAB外排泵组件的基因。在功能性磺胺类抗性筛选中，回收了folP基因，其编码突变的二氢蝶酸合酶，这是磺胺类药物作用的靶点。从功能性筛选中回收的基因来自共生菌的染色体，这些共生菌具有机会致病性，并且能够与相关致病菌种交换DNA。通过微阵列鉴定的基因在基于活性的筛选中未被回收，这表明这两种方法在促进鉴定人类微生物群中存在的一系列抗性机制方面可以互补。这也进一步证明了人类肠道和唾液中细菌群体中存在多种抗性机制。未来，本研究中描述的方法可用于监测抗生素治疗后抗性组的变化。
