Importantly, Kamal and Dennis recently showed that PAS is not affected by the antibiotic resistance status of the targeted cell [40]. Even for multi-antibiotic-resistant bacteria such as S. aureus, Burkholderia cenocepacia, or P. aeruginosa, several combinations of antibiotics and phages have proven successful [39–41]. Antibiotics belonging to different classes and having different mechanisms of action exhibited PAS when combined with phages (Table 1), compared to the few synergistic combinations reported for antibiotic cocktails. In addition, PAS works not only against bacteria in the planktonic phase but also in protective structures such as biofilms, due to the ability of certain phages to penetrate such structures [35,38]. Some bacterial morphological changes, such as filamentation and clustering, appear to facilitate the process of bacterial lysis resulting in increases in phage plaque size or phage titers [37,39,40]. In short, antibiotics can increase the efficacy of phages at eliminating bacterial populations, but the underlying mechanisms of this synergy with antibiotics remain uncertain and should be elucidated for the more refined application of PAS combination therapies (see Outstanding Questions).
重要的是，Kamal 和 Dennis 最近发现，PAS 不受目标细胞抗生素耐药性状态的影响 [40]。即使对于金黄色葡萄球菌、伯克霍尔德氏球菌或铜绿假单胞菌等多重抗生素耐药细菌，几种抗生素与噬菌体的组合也被证明是成功的[39-41]。属于不同类别、具有不同作用机制的抗生素在与噬菌体结合时都表现出 PAS 作用（表 1），而报道的抗生素鸡尾酒的协同作用组合却很少。此外，由于某些噬菌体具有穿透生物膜等保护性结构的能力，因此 PAS 不仅对浮游阶段的细菌有效，对生物膜等保护性结构中的细菌也有效 [35,38]。一些细菌形态学变化，如纤丝化和聚集，似乎有助于细菌裂解过程，从而导致噬菌体斑块大小或噬菌体滴度的增加 [37,39,40]。简而言之，抗生素可以提高噬菌体消灭细菌种群的效率，但这种与抗生素协同作用的潜在机制仍不确定，应在 PAS 组合疗法的更广泛应用中加以阐明（见未决问题）。
Rational phage–antibiotic combinations would appear impervious to the past failures of antibiotics. Still, the pharmacodynamics of antimicrobial combinations and dosing levels and schedules should also be incorporated into scientific approaches treating bacterial infections [48] (see Outstanding Questions). Evolutionary insights and approaches can help clinical researchers identify both inappropriate antimicrobial combinations and, when two or more are employed sequentially, orderings that amplify the selection of highly resistant bacterial populations [49]. Moreover, obviously, the use of lysogenic phages, with their limited impacts on bacterial populations and their capacity to transfer antibiotic-resistance genes has little potential for phage–antibiotic combined therapies.
合理的噬菌体-抗生素组合似乎不会受到过去抗生素失败的影响。不过，抗菌药组合的药效学以及剂量水平和时间表也应纳入治疗细菌感染的科学方法中[48]（见未决问题）。进化论的观点和方法可以帮助临床研究人员识别不恰当的抗菌药组合，以及在连续使用两种或两种以上抗菌药时，会扩大高耐药性细菌种群选择的排序[49]。此外，由于溶菌性噬菌体对细菌种群的影响有限，而且其转移抗生素耐药基因的能力也有限，因此使用溶菌性噬菌体进行噬菌体-抗生素联合疗法的可能性显然很小。
We see four possible stumbling blocks to phage–antibiotic combination therapies. First, strong selection for double-resistant variants in combination therapies is a serious concern. Recent work shows this for antibiotic cocktails [43] but, to our knowledge, there is no analogous experimental evidence for combined phage–antibiotic treatments.
我们认为噬菌体-抗生素联合疗法可能存在四个绊脚石。首先，在联合疗法中对双重抗性变体的强烈选择是一个严重问题。最近的研究表明鸡尾酒抗生素也会出现这种情况[43]，但据我们所知，噬菌体-抗生素联合疗法还没有类似的实验证据。
Potential Drawbacks of Phage–Antibiotic Combinations
噬菌体-抗生素组合的潜在缺点
Second, phages that preferentially target antibiotic-sensitive variants may promote antibiotic-resistant subpopulations through a phenomenon similar to ‘competitive release’, observed in antibiotic combinations [43]. According to this hypothetical scenario, previously antibiotic-treated bacteria can harbor sensitive subpopulations that could be preferentially targeted by phages and thus indirectly favor antibiotic-resistant variants. For example, phages may prefer antibiotic-sensitive active bacteria as hosts rather than persisters or bacteria forming antibiotic-resistant biofilms. Extensive work on antibiotic therapies and pathogenesis, and the use of biofilm-degrading phages or phage cocktails, limit this concern [30,35,38].
其次，优先针对抗生素敏感变体的噬菌体可能会通过一种类似于抗生素组合中观察到的 “竞争性释放 “现象来促进抗生素亚群的发展[43]。根据这种假设情况，以前用抗生素处理过的细菌中可能存在敏感亚群，而噬菌体可能会优先攻击这些亚群，从而间接促进抗生素耐药变体的产生。例如，噬菌体可能更倾向于选择对抗生素敏感的活性细菌作为宿主，而不是持久性细菌或形成抗生素生物膜的细菌。对抗生素疗法和致病机理的广泛研究，以及生物膜降解噬菌体或噬菌体鸡尾酒的使用，限制了这种担忧 [30,35,38]。
Third, although screening would eliminate from consideration phage–antibiotic combinations where either agent alone is better than both together, the effect of acceptable combinations may be less than additive, for example, if antibiotics damage hosts in a way that phage infection is aborted or does not produce relevant quantities of offspring or, conversely, if phages block the absorption of the antibiotic by the targeted cell (e.g., E. coli phages attaching to porins could potentially block the entry of certain antibiotics [50]). On the contrary, the general synergistic phenomenon observed suggests that it is not the case and that phages can better infect or produce a more dramatic reduction in bacterial populations when combined with antibiotics. Even in hypothetical combinations where the phage life cycle was partially inhibited by antibiotics or by antibiotic-resistant bacteria, sufficient genetic variation in the phage population could foster adaptation and augmented bacterial control.
第三，尽管筛选可以排除单独使用其中一种药剂比同时使用两种药剂效果更好的噬菌体-抗生素组合，但可接受的组合效果可能不具有相加性，例如，如果抗生素对宿主造成损害，导致噬菌体感染中止或不产生相关数量的后代，或者相反，如果噬菌体阻碍目标细胞对抗生素的吸收（例如，附着在孔蛋白上的大肠杆菌噬菌体可能会阻碍某些抗生素的进入[50]）。相反，观察到的普遍协同现象表明，情况并非如此，噬菌体与抗生素结合后能更好地感染细菌或更显著地减少细菌数量。即使在噬菌体生命周期受到抗生素或抗生素耐药菌部分抑制的假想组合中，噬菌体种群中足够多的遗传变异也能促进适应性并增强对细菌的控制。
Fourth, phages and antibiotics may each independently modulate bacterial virulence, either through plastic responses such as quorum sensing, or the competitive release of virulent variants (Box 1). Effects of combined therapy, such as reduced bacterial densities and increased costs, suggest that virulence would likely decrease, although there can be exceptions (Box 1).
第四，噬菌体和抗生素可通过法定人数感应等可塑性反应或毒性变体的竞争性释放，各自独立调节细菌的毒力（方框 1）。联合疗法的效果，如降低细菌密度和增加成本，表明毒力可能会降低，但也可能有例外（方框 1）。
Although there is little or no experimental evidence indicating these four phenomena as important obstacles, each will need to be properly evaluated in decisions on whether or not to combine phages and antibiotics in any given treatment.
尽管几乎没有实验证据表明这四种现象是重要障碍，但在决定是否在任何特定治疗中将噬菌体和抗生素结合使用时，需要对每种现象进行适当评估。
Box 1. Combined Therapy Effects on Virulence
方框 1.联合疗法对病毒性的影响
A potential undesirable consequence of phage–antibiotic therapies is the aggravation of an infection by producing virulent mutant bacterial strains that may worsen the patient’s outcome if associated with resistance, or be transmitted to other hosts. Although this potential concern is currently unexplored for combined applications, evidence for it from single antimicrobials is conflicting. For example, quorum sensing may coordinate the production of certain virulence factors [51], and exposure to phages or antibiotics individually has been shown to select cooperative variants with instances of increased virulence [52,53]. However, phages targeting quorum-sensing receptors could alleviate this type of interaction and the potential risk of increased virulence [54]. Antibiotic resistance has been often associated with virulence, as in a recent screening associating Pseudomonas aeruginosa resistance with a fitness advantage in vitro and during in vivo infection [55]. However the opposite relationship has also been observed, as in the case of the loss of porins related to both higher antibiotic resistance and attenuated virulence in Acinetobacter baumannii [56]. By contrast, a number of studies show that bacteria resistant to lytic phages have attenuated virulence on different hosts, from fish or plants to humans [57–59]. A possible explanation is that the costs of resistance to phages affect the expression of pathogenicity factors or decrease bacterial growth capacity, both resulting in less virulent bacterial variants [57–59].
噬菌体抗生素疗法的一个潜在不良后果是通过产生毒性突变细菌菌株加重感染，如果这些菌株具有抗药性，可能会加重患者的病情，或传染给其他宿主。虽然这种潜在的担忧目前还没有在联合应用中得到探讨，但单种抗菌药物的证据却令人信服。例如，法定人数感应可能会协调某些毒力因子的产生 [51]，而单独接触噬菌体或抗生素已被证明会选择毒力增强的合作变体 [52,53]。然而，以法定人数感应受体为靶标的噬菌体可以减轻这种相互作用以及毒力增强的潜在风险 [54]。抗生素耐药性往往与毒力有关，如最近的一项筛选发现，铜绿假单胞菌的耐药性与体外和体内感染时的毒性优势有关 [55]。不过，也观察到了相反的关系，如鲍曼不动杆菌（Acinetobacter baumannii）的孔蛋白缺失与抗生素耐药性增强和毒力减弱有关 [56]。相比之下，许多研究表明，对溶菌噬菌体有抗性的细菌对不同宿主（从鱼类或植物到人类）的毒力减弱 [57-59]。一种可能的解释是，对噬菌体产生抗性的代价会影响致病因子的表达或降低细菌的生长能力，从而导致细菌变种的毒性降低 [57-59]。
Outstanding Questions 未决问题
What are the underlying molecular evolutionary mechanisms of the synergistic interaction of antibiotic–phage cocktails?
抗生素-噬菌体鸡尾酒协同作用的分子进化机制是什么？
Can we assess optimum combinations of different types of phages and antibiotics that can then be extrapolated to treat other bacteria?
我们能否评估出不同类型噬菌体和抗生素的最佳组合，然后再推而广之，用于治疗其他细菌？
Which antibiotic doses achieve acceptable or optimal effects when combined with phages?
哪些抗生素剂量与噬菌体结合能达到可接受或最佳效果？
What is the best timing (sequential, simultaneous) and antimicrobial sequence (first phages then antibiotics or vice versa)?
什么是最佳时机（先后或同时）和抗菌顺序（先用噬菌体，再用抗生素，反之亦然）？
Can combined antibiotic and phage resistance emerge and what are the consequences for the patient and eventual transmission to other hosts?
是否会出现抗生素和噬菌体的联合耐药性，对患者以及最终传染给其他宿主的后果是什么？
What type of scientific evidence is needed on phage therapy to help satisfy safety regulations and legal constraints?
噬菌体疗法需要什么样的科学证据才能满足安全法规和法律约束的要求？
Concluding Remarks 结束语
Antibiotics have been extraordinarily successful at controlling bacterial pathogens. But horizontal gene transfer or de novo mutations resulting in increasing numbers of multidrug-resistant bacteria and the diminished discovery of new antimicrobial molecules lead to the unavoidable conclusion that other approaches are now necessary to conserve the action of existing molecules and maintain the high potency of future discoveries. Phages hold considerable potential, but we claim that achieving this will often mean combining them with antibiotics. The synergy observed between antibiotics and phages is a general phenomenon described in many studies, but not yet sufficiently understood and developed to reach actual application. We argue that evolutionary biology provides a framework for understanding control successes and failures of combined therapies, and how and whether we can adapt measures to specific situations. A number of unanswered questions regarding the use of phages as complements to antibiotics provide fertile ground for future research (see Outstanding Questions). Specifically, we need a deeper understanding of the molecular basis for combined antibiotic–phage therapies and in particular the basis for why antibiotic concentrations are so important in mediating outcomes. Our encouraging conclusion is that we have a second chance at controlling bacterial diseases and must avoid making the same mistakes as with the rampant use of antibiotics.
抗生素在控制细菌病原体方面取得了巨大成功。但是，横向基因转移或新基因突变导致耐多种药物的细菌数量不断增加，新抗菌分子的发现也越来越少，这就导致了一个不可避免的结论：现在必须采取其他方法来保护现有分子的作用，并保持未来发现的抗菌分子的高效力。噬菌体具有相当大的潜力，但我们认为，要实现这一目标，往往需要将噬菌体与抗生素结合起来。抗生素与噬菌体之间的协同作用是许多研究中描述的普遍现象，但尚未得到充分的理解和发展，以达到实际应用的目的。我们认为，进化生物学提供了一个框架，可用于理解联合疗法在控制方面的成功与失败，以及我们如何以及是否能根据具体情况调整措施。在使用噬菌体作为抗生素的补充方面，还有许多问题尚未解决，这为今后的研究提供了肥沃的土壤（见 “未决问题”）。具体来说，我们需要更深入地了解抗生素-噬菌体联合疗法的分子基础，尤其是抗生素浓度为何对治疗效果如此重要的基础。令人鼓舞的结论是，我们有第二次机会控制细菌性疾病，但必须避免重蹈抗生素肆意使用的覆辙。Post navigation