Bacteriophages, also known as phages, are viruses that exclusively target and infect bacterial cells [1]. The Caudalovirales, being the most extensively studied Order of phages, have an established structural arrangement. It comprises a protein capsid that encloses a double-stranded DNA genome, capped by a portal structure that links to a multi-component molecular machine called the tail. The tail functions first to interact with a receptor located on the surface of the host bacteria, thus initiating the process of infection, and thereafter functions as the conduit through which the phage DNA traverses to the bacterial cytoplasm [2]. Conceptually, the Caudalovirales are classified into three groupings based on their tails: the Siphoviridae (long-tailed, non-contractile phage), Myoviridae (long-tailed, contractile phage), and the Podoviridae (short-tailed, noncontractile phage) [3]. In 1936, a unique phage belonging to the Siphoviridae class was identified and shown to infect Salmonella enterica [4]. Named Chi (c) phage, it stood as something of a curiosity until the discovery of other Salmonella-targeting bacteriophage in waterways in the United Kingdom and new mechanistic insights into the recognition of bacterial flagellae by Chi-like phages [5]. A cryogenic electron microscopy (cryo-EM) study revealed the construction and stabilization of YSD1, a flagellotropic-tailed Chi-like Salmonella-targeting phage [6].
In our current study, we will present two novel species of Chi-like phages isolated in southern Australia, namely PIN1 phage and PIN2 phage. Comparative genomics analyses have demonstrated the presence of similar genomic areas across the four Chi-like phages (c, YSD1, PIN1, and PIN2). However, it is notable that these phages exhibit different variations in their tail fiber proteins. Further, the infection of Klebsiella by PIN1 and PIN2 requires the presence of an alternative receptor, given that Klebsiella lacks the capacity to produce flagella. Structural determination of novel Klebsiella-targeting phages will contribute to the advancement of our understanding of how Chi-like phages target their target bacterial host lacking flagella and may provide insight into targeting other non-flagellated bacterial species by their respective phages.