SGP-C: A Broad Host Range Temperate Bacteriophage; Against Salmonella gallinarum

Salmonella gallinarum is a poultry restricted-pathogen causing fowl-typhoid disease in adult birds with mortality rates up-to 80% and exhibit resistance against commonly used antibiotics. In this current study, a temperate broad host range bacteriophage SGP-C was isolated against S. gallinarum from poultry digesta. It showed infection ability in all the 15 tested field strains of S. gallinarum. The SGP-C phage produced circular, turbid plaques with alternate rings. Its optimum activity was observed at pH 7.0 and 37–42°C, with a latent period of 45 min and burst size of 187 virions/bacterial cell. The SGP-C lysogens, SGPC-L5 and SGPC-L6 exhibited super-infection immunity against the same phage, an already reported feature of lysogens. A virulence index of 0.5 and 0.001 as MV50 of SGP-C suggests its moderate virulence. The genome of SGP-C found circular double stranded DNA of 42 Kbp with 50.04% GC content, which encodes 63 ORFs. The presence of repressor gene at ORF49, and absence of tRNA sequence in SGP-C genome indicates its lysogenic nature. Furthermore, from NGS analysis of lysogens we propose that SGP-C genome might exist either as an episome, or both as integrated and temporary episome in the host cell and warrants further studies. Phylogenetic analysis revealed its similarity with Salmonella temperate phages belonging to family Siphoviridae. The encoded proteins by SGP-C genome have not showed homology with any known toxin and virulence factor. Although plenty of lytic bacteriophages against this pathogen are already reported, to our knowledge SGP-C is the first lysogenic phage against S. gallinarum reported so far.

Characterization of Phage Obtained from Methicillin -Resistant Staphylococcus aureus (MRSA)

Objective: The emergence of Staphylococcus aureus strains resistant to all antimicrobials and failure to discover new antibiotics have led researchers to phage therapy, which lost popularity after the discovery of antibiotics. The development of recombinant technology introduced the idea of creating lysogenic recombinant phages that provide controlled bacterial death and this required small- sized phages that were easy to manipulate. Our aim is to identify small-sized lysogenic bacteriophages that can be used safely in therapy. Method: The gene and protein map of the phage was created by analysis of sequencing after extracting a phage from the MRSA strain that is known to contain a small phage. Results: The phage was classified in Caudovirales spp. as it contains genes encoding tail proteins, and in Podoviridae spp. due to its genomic size and arrangement. Conclusion: To date, there are only sixteen phages from Podoviridae family uploaded on NCBI, and the phage described in this study is the seventeenth one. Only 41.4% of the ORFs (Open Reading Frames) in the genome could be matched with proteins using the NCBI BLAST. Recent studies suggest that 50-75% of bacteriophage ORFs do not correspond to any organism in GenBank. For better understanding of bacteriophages and their utilization in phage therapy, it is essential to sequence greater number of phages, and to discover their genomes and corresponding proteins. Since the genes and proteins of a lysogenic phage that can be used safely in recombinant phage therapies have been identified in our study, it will contribute to the relevant literature.

Lysogenization of a Lactococcal Host with Three Distinct Temperate Phages Provides Homologous and Heterologous Phage Resistance

Lactococcus lactis is the most widely exploited microorganism in global dairy fermentations. Lactococcal strains are described as typically harboring a number of prophages in their chromosomes. The presence of such prophages may provide both advantages and disadvantages to the carrying host. Here, we describe the deliberate generation of three distinct lysogens of the model lactococcal strain 3107 and the impact of additional prophage carriage on phage-resistance and anti-microbial susceptibility. Lysogen-specific responses were observed, highlighting the unique relationship and impact of each lysogenic phage on its host. Both homologous and heterologous phage-resistance profiles were observed, highlighting the presence of possible prophage-encoded phage-resistance factors. Superinfection exclusion was among the most notable causes of heterologous phage-resistance profiles with resistance observed against members of the Skunavirus, P335, P087, and 949 lactococcal phage groups. Through these analyses, it is now possible to identify phages that may pursue similar DNA injection pathways. The generated lysogenic strains exhibited increased sensitivity to the antimicrobial compounds, nisin and lysozyme, relative to the parent strain, although it is noteworthy that the degree of sensitivity was specific to the individual (pro)phages. Overall, the findings highlight the unique impact of each prophage on a given strain and the requirement for strain-level analysis when considering the implications of lysogeny.

Combined Use of the Ab105-2φΔCI Lytic Mutant Phage and Different Antibiotics in Clinical Isolates of Multi-Resistant Acinetobacter baumannii

Phage therapy is an abandoned antimicrobial therapy that has been resumed in recent years. In this study, we mutated a lysogenic phage from Acinetobacter baumannii into a lytic phage (Ab105-2phiΔCI) that displayed antimicrobial activity against A. baumannii clinical strain Ab177_GEIH-2000 (isolated in the GEIH-REIPI Spanish Multicenter A. baumannii Study II 2000/2010, Umbrella Genbank Bioproject PRJNA422585, and for which meropenem and imipenem MICs of respectively, 32 µg/mL, and 16 µg/mL were obtained). We observed an in vitro synergistic antimicrobial effect (reduction of 4 log–7 log CFU/mL) between meropenem and the lytic phage in all combinations analyzed (Ab105-2phiΔCI mutant at 0.1, 1 and 10 MOI and meropenem at 1/4 and 1/8 MIC). Moreover, bacterial growth was reduced by 8 log CFU/mL for the combination of imipenem at 1/4 MIC plus lytic phage (Ab105-2phiΔCI mutant) and by 4 log CFU/mL for the combination of imipenem at 1/8 MIC plus lytic phage (Ab105-2phiΔCI mutant) at both MOI 1 and 10. These results were confirmed in an in vivo model (G. mellonella), and the combination of imipenem and mutant Ab105-2phiΔCI was most effective (p < 0.05). This approach could help to reduce the emergence of phage resistant bacteria and restore sensitivity to antibiotics used to combat multi-resistant strains of Acinetobacter baumannii.

RNA-seq and two-dimensional gel electrophoresis revealed genomic and proteomic signatures during bacteriophage-host interactions in Pseudomonas aeruginosa

Background Understanding the biological nature of bacteriophage is important in exploring the therapeutic and biotechnological potentials of bacteriophages. However, available information is limited to the infection processes on either model phages infecting Escherichia coli or lytic phages against pathogens. The interplay between lysogenic phage and its host was rarely studied. Results We investigated the interactions between Pseudomonas aeruginosa and a lysogenic bacteriophage PaP3 through RNA-seq and two-dimensional gel electrophoresis (2D-GE). Compared to the uninfected host, a total of 2,891 (51.3%) differentially expressed genes (DGEs) were identified, most of which were repressed by phages, including the changes in metabolic-related and virulence-associated genes. The RT-qPCR results showed consistent directional changes compared with the RNA-seq results. According to 2D-GE, phage structure proteins were detected after phage infection. The host proteins, such as flagella hook-associated proteins, disappeared gradually after phage infection and may be shut off by phage. Conclusions All these indicate that although lysogenic phages do not immediately lyse the host, they play a significant regulatory role in the expression of host genes. Our findings provide an expanded view of the lysogenic phage infection processes and may offer potential targets for therapeutic intervention against P. aeruginosa infections.

Genomic Analysis of the Polyvalent Bacteriophage FP01

Recently the polyvalent bacteriophage FP01, isolated from wastewater in Valparaiso, Chile, was described to have lytic activity across species against Escherichia coli and Salmonella enterica serovars. Due to it polyvalent nature the bacteriophage FP01 could have potential application in food and agri-industry. Also, fundamental aspects of polyvalent bacteriophage biology are not well known. In this study we sequenced and describe the complete genome of the polyvalent phage FP01 (MH745368) using the nanopore technology. The bacteriophage FP01 genome has a 44,900 bp, double-stranded DNA with an average G+C content of 49.41% and 90 coding sequences (CDSs). We found that the phage FP01 critically depends on host factors for replication and transcription. Also, it has a critical lysogenic repressor pseudogene. Phylogenetic analyses indicated that the phage FP01 is closely related to phages lambda and P22. These results suggest that the phage FP01 could be a lytic variant of a lysogenic phage or acquired genes from lysogenic phages during host infection.

Complete Genome Sequence ofBacillus subtilisStrain CU1050, Which Is Sensitive to Phage SPβ

The Gram-positive bacteriumBacillus subtilisis used as a model organism to study cellular and molecular processes. Here, we announce the complete genomic sequence ofB. subtilisstrain CU1050, derived fromB. subtilisstrain 168. CU1050 has historically been used to study suppressor mutations and phage biology, especially the lysogenic phage SPβ.