Phage Lysogeny and Inhibition of Biofilm Formation

描述(申请人提供)：我们之前报道了一种铜绿假单胞菌转导噬菌体的分离和鉴定，命名为DMS3，最初从该菌的一种临床菌株中分离出来。我们实验室最近的研究发现了这种溶原性噬菌体的一个新角色--在所有体外测试条件下抑制表面附着群落(称为生物膜)的形成的能力。当DMS3裂解实验室菌株PA14或几个临床菌株时，会导致溶原菌在我们测试的所有条件下都不能形成生物膜，包括在多种碳源(葡萄糖、酪氨酸、LB、精氨酸、有机酸)和各种表面(如聚氯乙烯和聚苯乙烯塑料、玻璃)上生长，并且在不影响细菌生长的情况下生长，表明噬菌体DMS3的溶原性干扰了铜绿假单胞菌生物膜形成所需的中心途径。虽然我们的大部分研究都涉及噬菌体DMS3，但我们也从铜绿假单胞菌的临床分离株中分离出了几个额外的噬菌体，这些噬菌体也抑制了铜绿假单胞菌生物被膜的形成。初步证据表明，其中一种名为DMS577的噬菌体与DMS3截然不同。综上所述，这些数据表明，噬菌体DMS3和其他噬菌体可以调节铜绿假单胞菌启动表面附着生活方式的能力。 我们推测，噬菌体DMS3或DMS577的溶原性通过干扰生物膜形成途径的一个重要组成部分来阻止生物膜的形成。细菌与其噬菌体的相互作用是多微生物感染的显著例子，这些感染导致细菌获得新的毒力因子，进化出新的致病性状，并提高噬菌体裂解原微生物在广泛宿主中致病的能力。我们的数据表明，作为铜绿假单胞菌多菌感染的一部分，噬菌体DMS3和DMS577可能具有影响感染的性质和结果的能力，包括慢性化脓性中耳炎。 为了验证上述假设，我们为R21的应用提出了以下特定目标：特定目标I.确定抑制生物被膜形成所需的噬菌体DMS3编码功能。特定目的2.确定噬菌体溶源阻断生物被膜形成的步骤。特定目的III.测定噬菌体DMS577的DNA序列及其干扰生物膜形成的机制。

DESCRIPTION (provided by applicant): We previously reported the isolation and characterization of a transducing phage of P. aeruginosa, designated DMS3, originally isolated from a clinical strain of this organism. Recent studies in our lab have uncovered a new role for this lysogenic bacteriophage - the ability to inhibit the formation of surface- attached communities (known as biofilms) under all in vitro conditions tested. DMS3, when lysogenizing the laboratory strain P. aeruginosa PA14 or several clinical strains, results in a lysogen unable to make a biofilm under every condition we have tested, including growth on a variety of carbon sources (glucose, casamino acids, LB, arginine, organic acids) and on a variety of surfaces (i.e., PVC & polystyrene plastic, glass), and does so without affecting bacterial growth, indicating that lysogeny by phage DMS3 interferes with a central pathway required for biofilm formation by P. aeruginosa. While most of our studies have involved phage DMS3, we also have isolated several additional phages from clinical isolates of P. aeruginosa that also inhibit biofilm formation by P. aeruginosa. Preliminary evidence suggests that one such phage, designated DMS577, is distinct from DMS3. Taken together, these data indicate that phage DMS3 and other phages can modulate the ability of P. aeruginosa to initiate a surface-attached lifestyle. We hypothesize that lysogeny by phage DMS3 or DMS577 blocks biofilm formation by interfering with an essential component of the biofilm formation pathway. The interaction of bacteria and their phages are striking examples of polymicrobial infections resulting in the bacterium acquiring new virulence factors, evolving new pathogenic traits and increasing the ability of the phage-lysogenized microbe to cause disease in a wide range of hosts. Our data suggest that as part of a polymicrobial infection with P. aeruginosa, bacteriophages DMS3 and DMS577 may have the ability to influence the nature and outcome of infections, including chronic suppurative otitis media. To test the hypotheses above, we propose the following Specific Aims for this R21 application: Specific Aim I. Determine the phage DMS3-encoded functions required to inhibit biofilm formation. Specific Aim 2. Determine the step in biofilm formation blocked by phage lysogeny. Specific Aim III. Determine DMA sequence of phage DMS577 and the mechanism by which DMS577 interferes with biofilm formation.