中文摘要：越来越多的抗生素耐药性问题引起了水产养殖业对噬菌体疗法的兴趣。然而，目标细菌的噬菌体耐药性演变是一个挑战。为了研究噬菌体耐药性如何影响鱼类病原体柱状黄杆菌，本研究将两种野生型细菌菌株FCO-F2和FCO-F9暴露于噬菌体（FCO-F2暴露于FCOV-F2、FCOV-F5和FCOV-F25，以及FCO-F9暴露于FCL-2、FCOV-F13和FCOV-F45），并分析由此产生的细菌表型和遗传变化。细菌活力在接触培养物后先降低，但在1至2天后开始增加，同时菌落形态从原始根状变为粗糙，导致粗糙形态类型的感染率为98%。在虹鳟鱼苗中，对24个分离株（包括4个来自无噬菌体处理的分离株）的噬菌体耐药性、抗生素敏感性、运动性、粘附性、生物膜形成、蛋白酶活性、全基因组测序和毒力进行了进一步鉴定。研究发现，在噬菌体暴露中产生的粗糙菌株具有噬菌体耐药性及低毒力，而根状菌株则保持了噬菌体敏感性和高毒力；滑动运动和蛋白酶活性也与噬菌体敏感性有关。噬菌体耐药菌株的突变大多位于编码IX型分泌系统的基因中，该系统是类杆菌滑动运动机制的组成部分。然而，并非所有的噬菌体耐药菌株都有突变，表明柱状黄杆菌的噬菌体耐药性是一个多因素的过程，包括基因突变和基因表达的变化。可见，噬菌体耐药性可能不是噬菌体治疗柱状黄杆菌感染的挑战，因为噬菌体耐药性关系到细菌毒力的降低。
重点：感染性细菌的噬菌体耐药性是一种常见现象，对噬菌体治疗的发展提出了挑战。随着世界人口的增长和对粮食产量需求的增加，不断加强的畜牧业面临着越来越多的传染病问题。由柱状黄杆菌引起的柱状病对鲑鱼鱼苗和幼鱼养殖构成了世界性威胁。如果不进行抗生素治疗，感染可致鱼类100%的死亡。用噬菌体治疗柱状病会降低水产养殖业耐抗生素细菌的发展和抗生素负荷，但耐噬菌体菌株可能成为一个新风险。不过，本研究中噬菌体抗性柱状黄杆菌分离株的表型和遗传特征显示，它们的毒性低于噬菌体敏感株，因此对噬菌体治疗柱状病并不构成威胁。可见，本文用噬菌体预防和治疗柱状黄杆菌感染的方法，对全球养鱼业很有参考价值。
外文摘要：Increasing problems with antibiotic resistance have directed interest toward phage therapy in the aquaculture industry. However, phage resistance evolving in target bacteria is considered a challenge. To investigate how phage resistance influences the fish pathogen Flavobacterium columnare, two wild-type bacterial isolates, FCO-F2 and FCO-F9, were exposed to phages (FCO-F2 to FCOV-F2, FCOV-F5, and FCOV-F25, and FCO-F9 to FCL-2, FCOV-F13, and FCOV-F45), and resulting phenotypic and genetic changes in bacteria were analyzed. Bacterial viability first decreased in the exposure cultures but started to increase after 1 to 2 days, along with a change in colony morphology from original rhizoid to rough, leading to 98% prevalence of the rough morphotype. Twenty-four isolates (including four isolates from no-phage treatments) were further characterized for phage resistance, antibiotic susceptibility, motility, adhesion, and biofilm formation, protease activity, whole-genome sequencing, and virulence in rainbow trout fry. The rough isolates arising in phage exposure were phage resistant with low virulence, whereas rhizoid isolates maintained phage susceptibility and high virulence. Gliding motility and protease activity were also related to the phage susceptibility. Observed mutations in phage-resistant isolates were mostly located in genes encoding the type IX secretion system, a component of the Bacteroidetes gliding motility machinery. However, not all phage-resistant isolates had mutations, indicating that phage resistance in F. columnare is a multifactorial process, including both genetic mutations and changes in gene expression. Phage resistance may not, however, be a challenge for development of phage therapy against F. columnare infections since phage resistance is associated with decreases in bacterial virulence.
   IMPORTANCE Phage resistance of infectious bacteria is a common phenomenon posing challenges for the development of phage therapy. Along with a growing world population and the need for increased food production, constantly intensifying animal farming has to face increasing problems of infectious diseases. Columnaris disease, caused by Flavobacterium columnare, is a worldwide threat for salmonid fry and juvenile farming. Without antibiotic treatments, infections can lead to 100% mortality in a fish stock. Phage therapy of columnaris disease would reduce the development of antibiotic-resistant bacteria and antibiotic loads by the aquaculture industry, but phage-resistant bacterial isolates may become a risk. However, phenotypic and genetic characterization of phage-resistant F. columnare isolates in this study revealed that they are less virulent than phage-susceptible isolates and thus not a challenge for phage therapy against columnaris disease. This is valuable information for the fish farming industry globally when considering phage-based prevention and curing methods for F. columnare infections.
 
外文关键词：bacteriophage；colony morphology；Flavobacterium columnare；gliding motility; mutation；phage resistance；type IX secretion system；virulence
作者：Kunttu, HMT；Runtuvuori-Salmela, A；Sundell, K；Wiklund, T；Middelboe, M；Landor, L；Ashrafi, R；Hoikkala, V；Sundberg, LR
作者单位：Univ Jyvaskyla；Abo Akad Univ；Univ Copenhagen；Univ Bergen
期刊名称：APPLIED AND ENVIRONMENTAL MICROBIOLOGY
期刊影响因子：4.016
出版年份：2021
出版刊次：16
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