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We report the discovery of a satellite-helper phage system with a novel type of dependence on a tail donor. The Acinetobacter baumannii satellite podovirus Aci01-2-Phanie (short name Phanie) uses a phage phi29-like DNA replication and packaging mode. Its linear 11,885 bp dsDNA genome bears 171 bp inverted terminal repeats (ITR). Phanie is related to phage DU-PP-III from Pectobacterium and to members of the Astrithrvirus from Salmonella enterica . Together, they form a new clade of phages with 27% to 30% identity over the whole genome. Detailed 3D protein structure prediction and mass spectrometry analyses demonstrate that Phanie encodes its capsid structural genes and genes necessary to form a short tail. However, our study reveals that Phanie virions are non-infectious unless they associate with the contractile tail of an unrelated phage, Aci01-1, to produce chimeric myoviruses. Following the coinfection of Phanie with myovirus Aci01-1, hybrid viral particles composed of Phanie capsids and Aci01-1 contractile tails are assembled together with Phanie and Aci01-1 particles.IMPORTANCEThere are few reported cases of satellite-helper phage interactions but many more may be yet undiscovered. Here we describe a new mode of satellite phage dependence on a helper phage. Phanie, like phage phi29, replicates its linear dsDNA by a protein primed-mechanism and protects it inside podovirus-like particles. However, these particles are defective, requiring the acquisition of the tail from a myovirus helper for production of infectious virions. The formation of chimeras between a phi29-like podovirus and a helper contractile tail reveals an unexpected association between very different bacterial viruses., Competing Interests: The authors declare no conflict of interest.

The Acinetobacter baumannii bacteriophage Aci01-1, which belongs to the genus Saclayvirus of the order Caudoviricetes, has an icosahedral head and a contractile rigid tail. We report that Aci01-1 has, attached to the tail conical tip, a remarkable 146-nm-long flexible fiber with seven beads and a terminal knot. Its putative gene coding for a 241.36-kDa tail fiber protein is homologous to genes in Aci01-1-related and unrelated phages. Analysis of its 3D structure using AlphaFold provides a structural model for the fiber observed by electron microscopy. We also identified a putative receptor of the phage on the bacterial capsule that is hypothesized to interact with the Aci01-1 long fiber., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Sixteen bacteriophages of Achromobacter xylosoxidans distributed into four genera have been isolated from sewage water in Abidjan, Côte d'Ivoire, using a single clinical strain, and their genomes have been sequenced. Three podoviruses belonged to the genus Phikmvvirus, and these represent the first A. xylosoxidans phages of this genus. Seven podoviruses, distributed into three groups, belonged to the genus Jwalphavirus. Among the siphoviruses, three revealed similarities to Pseudomonas phage 73 and members of the genus Septimatrevirus, and three were YuA-like phages. The virulence of these phages toward a panel of 10 genetically diverse strains was tested, with the phiKMV-like phages showing the broadest host range.

Phage therapy may become a complement to antibiotics in the treatment of chronic Pseudomonas aeruginosa infection. To design efficient therapeutic cocktails, the genetic diversity of the species and the spectrum of susceptibility to bacteriophages must be investigated. Bacterial strains showing high levels of phage resistance need to be identified in order to decipher the underlying mechanisms. Here we have selected genetically diverse P. aeruginosa strains from cystic fibrosis patients and tested their susceptibility to a large collection of phages. Based on plaque morphology and restriction profiles, six different phages were purified from "pyophage", a commercial cocktail directed against five different bacterial species, including P. aeruginosa. Characterization of these phages by electron microscopy and sequencing of genome fragments showed that they belong to 4 different genera. Among 47 P. aeruginosa strains, 13 were not lysed by any of the isolated phages individually or by pyophage. We isolated two new phages that could lyse some of these strains, and their genomes were sequenced. The presence/absence of a CRISPR-Cas system (Clustered Regularly Interspaced Short Palindromic Repeats and Crisper associated genes) was investigated to evaluate the role of the system in phage resistance. Altogether, the results show that some P. aeruginosa strains cannot support the growth of any of the tested phages belonging to 5 different genera, and suggest that the CRISPR-Cas system is not a major defence mechanism against these lytic phages.

The objective of this study was to determine the genetic diversity of multidrug-resistant (MDR) Pseudomonas aeruginosa strains isolated over a period of 12 months in two French hospitals and to test their susceptibility to bacteriophages. A total of 47 MDR isolates recovered from hospitalized patients were genotyped using multiple-locus variable number of tandem repeats analysis. The genotypes were distributed into five clones (including 19, 5, 5, 3, and 3 isolates, respectively) and 12 singletons. Comparison to 77 MDR strains from three other countries, and MLST analysis of selected isolates showed the predominance of international MDR clones. The larger clone, CC235, contained 59 isolates displaying different antibiotic resistance mechanisms, including the presence of the GES1, VIM-2, VIM-4, and IMP-1 β-lactamases. Three newly isolated P. aeruginosa bacteriophages were found to lyse 42 of the 44 analyzed strains, distributed into the different clonal complexes. This pilot study suggests that systematic genotyping of P. aeruginosa MDR strains could improve our epidemiological understanding of transmission at both the local (hospital) and the national level and that phage therapy could be an alternative or a complementary treatment to antibiotics for treating MDR-infected patients.

A novel temperate bacteriophage of Pseudomonas aeruginosa, phage vB_PaeP_Tr60_Ab31 (alias Ab31) is described. Its genome is composed of structural genes related to those of lytic P. putida phage AF, and regulatory genes similar to those of temperate phage PAJU2. The virion structure resembles that of phage AF and other lytic Podoviridae (S. enterica Epsilon 15 and E. coli phiv10) with similar tail spikes. Ab31 was able to infect P. aeruginosa strain PA14 and two genetically related strains called Tr60 and Tr162, out of 35 diverse strains from cystic fibrosis patients. Analysis of resistant host variants revealed different phenotypes, including induction of pigment and alginate overproduction. Whole genome sequencing of resistant variants highlighted the existence of a large deletion of 234 kbp in two strains, encompassing a cluster of genes required for the production of CupA fimbriae. Stable lysogens formed by Ab31 in strain Tr60, permitted the identification of the insertion site. During colonization of the lung in cystic fibrosis patients, P. aeruginosa adapts by modifying its genome. We suggest that bacteriophages such as Ab31 may play an important role in this adaptation by selecting for bacterial characteristics that favor persistence of bacteria in the lung. [ABSTRACT FROM AUTHOR]

Phage therapy may become a complement to antibiotics in the treatment of chronic Pseudomonas aeruginosa infection. To design efficient therapeutic cocktails, the genetic diversity of the species and the spectrum of susceptibility to bacteriophages must be investigated. Bacterial strains showing high levels of phage resistance need to be identified in order to decipher the underlying mechanisms. Here we have selected genetically diverse P. aeruginosa strains from cystic fibrosis patients and tested their susceptibility to a large collection of phages. Based on plaque morphology and restriction profiles, six different phages were purified from “pyophage”, a commercial cocktail directed against five different bacterial species, including P. aeruginosa. Characterization of these phages by electron microscopy and sequencing of genome fragments showed that they belong to 4 different genera. Among 47 P. aeruginosa strains, 13 were not lysed by any of the isolated phages individually or by pyophage. We isolated two new phages that could lyse some of these strains, and their genomes were sequenced. The presence/absence of a CRISPR-Cas system (Clustered Regularly Interspaced Short Palindromic Repeats and Crisper associated genes) was investigated to evaluate the role of the system in phage resistance. Altogether, the results show that some P. aeruginosa strains cannot support the growth of any of the tested phages belonging to 5 different genera, and suggest that the CRISPR-Cas system is not a major defence mechanism against these lytic phages. [ABSTRACT FROM AUTHOR]