An active beta-lactamase is a part of an orchestrated cell wall stress resistance network of Bacillus subtilis and related rhizosphere species

A hallmark of the Gram-positive bacteria, such as the soil-dwelling bacterium Bacillus subtilis, is their cell wall. Here, we report that d-leucine and flavomycin, biofilm inhibitors targeting the cell wall, activate the beta-lactamase PenP. This beta-lactamase contributes to ampicillin resistance in B. subtilis under all conditions tested. In contrast, both Spo0A, a master regulator of nutritional stress, and the general cell wall stress response, differentially contribute to beta-lactam resistance under different conditions. To test whether beta-lactam resistance and beta-lactamase genes are widespread in other Bacilli, we isolated Bacillus species from undisturbed soils, and found that their genomes can encode up to five beta-lactamases with differentiated activity spectra. Surprisingly, the activity of environmental beta-lactamases and PenP, as well as the general stress response, resulted in a similarly reduced lag phase of the culture in the presence of beta-lactam antibiotics, with little or no impact on the logarithmic growth rate. The length of the lag phase may determine the outcome of the competition between beta-lactams and beta-lactamases producers. Overall, our work suggests that antibiotic resistance genes in B. subtilis and related species are ancient and widespread, and could be selected by interspecies competition in undisturbed soils.

革兰氏阳性菌（Gram-positive bacteria）的标志性特征为其细胞壁，以土壤栖息细菌枯草芽孢杆菌（Bacillus subtilis）为例。本研究发现，靶向细胞壁的生物膜抑制剂D-亮氨酸（d-leucine）与黄霉素（flavomycin）可激活β-内酰胺酶（beta-lactamase）PenP。该β-内酰胺酶在所有受试条件下，均可赋予枯草芽孢杆菌氨苄青霉素（ampicillin）抗性。与之相对，营养胁迫的主调控因子Spo0A以及通用细胞壁应激反应，在不同条件下对β-内酰胺抗性的贡献存在显著差异。为验证β-内酰胺抗性与β-内酰胺酶基因在其他芽孢杆菌属（Bacilli）物种中是否广泛存在，我们从未受干扰的土壤中分离得到芽孢杆菌物种，发现其基因组最多可编码五种具有差异化活性谱的β-内酰胺酶。令人意外的是，环境来源β-内酰胺酶与PenP的活性，以及通用应激反应，均可使培养物在β-内酰胺类抗生素存在下的延滞期显著缩短，且对对数生长速率几乎无影响。延滞期的长短或可决定β-内酰胺类抗生素与β-内酰胺酶产生菌之间的竞争结局。综上，本研究表明枯草芽孢杆菌及其近缘物种中的抗生素抗性基因古老且分布广泛，其可能在未受干扰的土壤中通过种间竞争被选择保留。