The putative drug efflux systems of the Bacillus cereus group

The Bacillus cereus group of bacteria includes seven closely related species, three of which, B. anthracis, B. cereus and B. thuringiensis, are pathogens of humans, animals and/or insects. Preliminary investigations into the transport capabilities of different bacterial lineages suggested that genes encoding putative efflux systems were unusually abundant in the B. cereus group compared to other bacteria. To explore the drug efflux potential of the B. cereus group all putative efflux systems were identified in the genomes of prototypical strains of B. cereus, B. anthracis and B. thuringiensis using our Transporter Automated Annotation Pipeline. More than 90 putative drug efflux systems were found within each of these strains, accounting for up to 2.7% of their protein coding potential. Comparative analyses demonstrated that the efflux systems are highly conserved between these species; 70–80% of the putative efflux pumps were shared between all three strains studied. Furthermore, 82% of the putative efflux system proteins encoded by the prototypical B. cereus strain ATCC 14579 (type strain) were found to be conserved in at least 80% of 169 B. cereus group strains that have high quality genome sequences available. However, only a handful of these efflux pumps have been functionally characterized. Deletion of individual efflux pump genes from B. cereus typically had little impact to drug resistance phenotypes or the general fitness of the strains, possibly because of the large numbers of alternative efflux systems that may have overlapping substrate specificities. Therefore, to gain insight into the possible transport functions of efflux systems in B. cereus, we undertook large-scale qRT-PCR analyses of efflux pump gene expression following drug shocks and other stress treatments. Clustering of gene expression changes identified several groups of similarly regulated systems that may have overlapping drug resistance functions. In this article we review current knowledge of the small molecule efflux pumps encoded by the B. cereus group and suggest the likely functions of numerous uncharacterised pumps.蜡样芽孢杆菌群（Bacillus cereus group）包含7个亲缘关系密切的菌种，其中炭疽芽孢杆菌（Bacillus anthracis）、蜡样芽孢杆菌（Bacillus cereus）与苏云金芽孢杆菌（Bacillus thuringiensis）3种可引发人类、动物及/或昆虫感染致病。针对不同细菌谱系的转运能力开展的初步研究显示，相较于其他细菌类群，蜡样芽孢杆菌群中编码推定外排系统的基因数量异常丰富。为探究蜡样芽孢杆菌群的药物外排潜力，本研究依托自主开发的转运蛋白自动注释流程（Transporter Automated Annotation Pipeline），在蜡样芽孢杆菌、炭疽芽孢杆菌与苏云金芽孢杆菌的模式菌株基因组中，完成了全部推定外排系统的鉴定工作。上述各模式菌株中均鉴定出90余种推定药物外排系统，占其编码蛋白总潜能的最高可达2.7%。比较分析结果显示，该三类菌种的外排系统高度保守：所研究的三株菌株间共有70%~80%的推定外排泵。此外，蜡样芽孢杆菌模式菌株ATCC 14579（即该菌种的标准模式菌株）所编码的推定外排系统蛋白中，有82%在169株拥有高质量基因组序列的蜡样芽孢杆菌群菌株中，至少80%的菌株内存在保守同源序列。然而，目前仅对其中极少数的外排泵完成了功能表征。在蜡样芽孢杆菌中单独敲除外排泵基因，通常对其耐药表型及菌株整体生存适合度几乎无显著影响，这可能是由于存在大量底物特异性存在重叠的替代外排系统。因此，为深入解析蜡样芽孢杆菌群外排系统可能的转运功能，本研究针对药物冲击及其他应激处理后的外排泵基因表达情况，开展了大规模实时定量逆转录聚合酶链反应（quantitative real-time reverse transcription polymerase chain reaction，简称qRT-PCR）分析。对基因表达变化模式进行聚类分析后，本研究鉴定出多组调控模式相似的外排系统，这类系统可能具备重叠的药物耐药功能。本文综述了当前关于蜡样芽孢杆菌群所编码的小分子外排泵的研究认知，并对大量尚未完成功能表征的外排泵的潜在功能进行了推测。