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The most frequent infections caused by Pseudomonas aeruginosa are local infections in soft tissues, including burns. Today, phage use is considered a suitable alternative to cure infections caused by multi-drug-resistant (MDR) and extensively drug-resistant (XDR) bacteria. We investigated the potential of a novel phage (vB_PaS-HSN4) belonging to Caudoviricetes class, against XDR and MDR P. aeruginosa strains in vivo and in vitro. Its biological and genetic characteristics were investigated. The phage burst size and latent were 119 and 20 min, respectively. It could tolerate a broad range of salt concentrations, pH values, and temperatures. The combination with ciprofloxacin significantly enhanced biofilm removal after 24 h. The genome was dsDNA with a size of 44,534 bp and encoded 61 ORFs with 3 tRNA and 5 promoters. No virulence factor was observed in the phage genome. In the in vivo infection model, treatment with vB_PaS-HSN4 increased Galleria mellonella larvae survival (80%, 66%, and 60%) (MOI 100) and (60%, 40%, and 26%) (MOI 1) in the pre-treatment, co-treatment, and post-treatment experiments, respectively. Based on these characteristics, it can be considered for the cure of infections of burns caused by P. aeruginosa., (© 2024. The Author(s).)

Periodontitis is a chronic inflammatory condition that can damage soft tissues and supporting teeth. Enterococcus faecalis is an opportunistic pathogen usually living in the oral cavity and plays a critical role in apical periodontitis that significantly threatens human health. The use of bacteriophages as an alternative way to eliminate bacterial infections is a promising approach. E. faecalis was isolated from the depth of dental packets of patients with periodontitis. Antimicrobial susceptibility was tested using 16 antimicrobial agents. Also, a specific virulent bacteriophage (vB_EfaS-SRH2) with an irregular pentagonal morphology of the head and a non-contractile tail belonging to the Siphoviridae, was isolated from wastewater in East of Isfahan, Iran, and its physiological and genomic specifications were investigated. The genome was double-strand DNA with 38,746 bp length and encoded 62 putative ORFs. In addition, eight Anti-CRISPERs and 30 Rho-dependent terminators were found. No tRNA was found. It had a short latent period of 15 min and a large burst size of ~ 125. No undesirable genes (antibiotic resistance, lysogenic dependence, and virulence factors) were identified in the genome. Based on physiological properties and genomic characteristics, this phage can be used as a suitable choice in phage therapy for periodontitis and root canal infection., (© 2022. The Author(s).)

The bacteriophage (phage) DNA extraction methods for genomics analysis is a critical and time-consuming process. Hence, a rapid and cost-effective method for DNA extraction of phages is favorable for phage biologists. In the present study, a cost-effective, simple and rapid procedure for phage genome extraction in less than 10 min is introduced. Highly concentrated phage lysates were prepared using acetone precipitation followed by extraction using various methods such as commercial kits, TES lysis buffer, potassium iodide, and sodium iodide. The quality of the extracted DNA was analyzed by agarose gel electrophoresis and UV absorbance of DNA at 260 and 280 nm. Finally, the extracted DNA was subjected to restriction digestion and next-generation sequencing to approve the efficiency of the method. Based on the time, cost, and quality of obtained DNA, the acetone precipitation of phages and extraction by potassium iodide or sodium iodide method was determined to be the best method for phage DNA extraction tested in this study. Moreover, the extracted genomic DNA using this method is suitable for phage genomic analysis such as restriction enzyme studies, preparation of DNA library, and also next-generation sequencing., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Enterococcus faecalis is an environmental agent of bovine mastitis in cows and has many cytopathic effects on the urinary tract in both humans and animals. In this study, a novel lytic bacteriophage, vB_EfaS-DELF1, was isolated against 21 E. faecalis isolated from bovine mastitis, including vancomycin-resistant E. faecalis (VRE). vB_EfaS-DELF1 bacteriophage was specific for E. faecalis and showed no lytic effects against other tested Enterococcus spp., Gram-negative, or Gram-positive bacteria. Moreover, no activity was observed against yogurt starters. The phage suspension was stable in a wide range of pH, salinity, and temperature. It retained its activity in 3.5 % fat milk. vB_EfaS-DELF1 has the common phenotypic features of Siphoviridae with a double-strand DNA of 40,248 bp in length and a G + C content of 34.9 %. The genome encodes 62 putative ORFs and no tRNA. No undesirable genes such as lysogenic mediators, antibiotic resistance, or virulence factor genes were detected in the genome. The comparative genomic analysis demonstrated similarity to the other available phage genomes. The highest similarity was observed with two other phages (50 % coverage and 82.38 % identities with IME-EFm1; 35 % coverage and 86.22 % identities with IME-EFm5) that were placed in the same clade. The differences with the other aligned phages were high and were placed in distant clusters. Regarding the specificity of this new bacteriophage against all of the tested E. faecalis isolates and, in particular, against the vancomycin-resistant ones, and also the absence of antibiotic resistance or virulence genes in its genome, vB_EfaS-DELF1 is suggested as a potential candidate for biocontrol of E. faecalis infections., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Shigella spp. can be isolated from various food sources and is responsible for many outbreaks and sporadic cases of foodborne diseases worldwide. Although Shigella species are known as one of the major foodborne pathogens, a few studies have characterized the prevalence and molecular basis of antibiotic resistance of Shigella spp. isolated from food origins. This study investigated the prevalence of Shigella spp. in a wide range of food samples (1400 samples), and the phenotypic and genotypic basis of antimicrobial resistance of the isolates. In addition, the potential of two Shigella specific phages (vB_SflS-ISF001 and vB_SsoS-ISF002) to control the growth of the isolates in food was tested. Shigella sonnei and Shigella flexneri were detected in 11 (0.8%) and 8 (0.6%) samples, respectively. The highest prevalence of Shigella spp. was observed in vegetables. Multidrug resistance phenotypes were noticeably frequent and observed in 17 isolates (89.5%) out of 19 isolates. Moreover, 13 (68.4%), 9 (47.4%) and 17 (89.5%) isolates were positive for β-lactamase-encoding, plasmid-mediated quinolone resistance and tetracycline resistance genes, respectively. Treatment with the phages reduced bacterial counts up to 3 and 4 log when used individually or in cocktail form, respectively. The findings of this study indicate the prevalence of Shigella spp. in food sources and also provide useful information for a better understanding of the molecular aspects of antimicrobial resistance in Shigella spp.. The results also suggest that the combination of vB_SflS-ISF001 and vB_SsoS-ISF002 phages can effectively reduce contamination of two important species of Shigella in food., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Bacteria of the genus Raoultella are known to inhabit aquatic environments and can be found in medical samples. The pathogenicity of Raoultella ornithinolytica isolates in human has become increasingly important, and several cases of infections have been reported recently. However, there are no reports of isolation of bacteriophages infecting this bacterium. In this study, two novel phages (ISF3 and ISF6) of a methylotrophic Raoultella strain were isolated from sewage. To characterize the isolated phages, morphological features, protein profiles, restriction digestion patterns, and partial genome sequences were studied. Despite morphological differences, electron microscopy revealed that both phages had an icosahedral capsid connected to a contractile tail, suggesting that ISF3 and ISF6 both belong to the family Myoviridae. Partial nucleotide sequences of the ISF3 genome showed 99% to 100% identity to DNA of Klebsiella pneumonia phages KP15, KP27 and BMBT1; however, the restriction digestion profiles of ISF3 genome digested by EcoRI and EcoRV differed from those of Klebsiella phages KP15 and KP27. A partial sequence alignment showed that ISF6 can be classified as a member of a new viral genus due to its significant differences from other previously identified phages. To the best of our knowledge, this is the first report of the isolation and characterization of the specific Raoultella phages that have potential to be used as new pharmaceuticals against R. ornithinolytica.

Proteus mirabilis is one of the most common causes of urinary tract infection (UTI), particularly in patients undergoing long-term catheterization. Phage vB_PmiS-TH was isolated from wastewater with high lytic activity against P. mirabilis (TH) isolated from UTI. The phage had rapid adsorption, a large burst size (∼260 PFU per infected cell), and high stability at a wide range of temperatures and pH values. As analyzed by transmission electron microscopy, phage vB_PmiS-TH had an icosahedral head of ∼87 × 62 nm with a noncontractile tail about 137 nm in length and 11 nm in width. It belongs to the family Siphoviridae. Combination of the phage vB_PmiS-TH with ampicillin had a higher removal activity against planktonic cells of P. mirabilis (TH) than the phage or the antibiotic alone. Combination of the phage at a multiplicity of infection of 100 with a high dose of ampicillin (246 µg/mL) showed the highest biofilm removal activity after 24 h. This study demonstrates that using a combination of phage and antibiotic could be significantly more effective against planktonic and biofilm forms of P. mirabilis (TH)., (© 2018 S. Karger AG, Basel.)

One of the most economically important bacterial pathogens of plants and plant products is Dickeya dadantii. This bacterium causes soft rot disease in tubers and other parts of the potato and other plants of the Solanaceae family. The application of restricted host range bacteriophages as biocontrol agents has recently gained widespread interest. This study purposed to isolate the infectious agent of the potato and evaluate its biocontrol by bacteriophages. Two phytopathogenic strains were isolated from infected potatoes, identified based on biochemical and 16S rRNA gene sequencing, and submitted to GenBank as D. dadantii strain pis3 (accession no. HQ423668) and D. dadantii strain sip4 (accession no. HQ423669). Their bacteriophages were isolated from Caspian Sea water by enriching the water filtrate with D. dadantii strains as hosts using spot or overlay methods. On the basis of morphotypes, the isolated bacteriophages were identified as members of the Myoviridae and Siphoviridae families and could inhibit the growth of antibiotic resistant D. dadantii strains in culture medium. Moreover, in Dickeya infected plants treated with bacteriophage, no disease progression was detected. No significant difference was seen between phage-treated and control plants. Thus, isolated bacteriophages can be suggested for the biocontrol of plant disease caused by Dickeya strains.

To date, a few numbers of bacteriophages that infect Lactococcus garvieae have been identified, but their complete genome sequences have not yet been investigated. For the first time, herein, the complete DNA sequence of a new phage of L. garvieae (phage WP-2) is reported and analyzed. The morphological characteristics indicated that the phage had a small isometric head along with a short and non-contractile tail, suggesting that WP-2 belongs to the family Podoviridae. Bioinformatic analysis revealed that phage WP-2 can be classified as a new member of Ahjdlikevirus in the Picovirinae subfamily because it had a small dsDNA of 18,899 bp with 24 open reading frames and a protein-primed DNA polymerase. The phage nucleotide sequence and predicted protein products have been identified to share very limited evidence of homology with complete genome and proteome of other phages. To our knowledge, this is the first Ahjdlikevirus bacteriophage which can infect a member of the Lactococcus genus., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Lactococcus garvieae is an emerging pathogen responsible for lactococcosis, a serious disease in trout aquaculture. The identification of new bacteriophages against L. garvieae strains may be an effective way to fight this disease and to study the pathogen's biology. Three L. garvieae phages, termed WP-1, WWP-2 and SP-2, were isolated from different environments, and their morphological features, genome restriction profiles and structural protein patterns were studied. Random cloning of HindIII-cut fragments was performed, and the fragments were partially sequenced for each phage. Although slight differences were observed by transmission electron microscopy, all of the phages had hexagonal heads and short non-contractile tails and were classified as members of the family Podoviridae. Restriction digestion analysis of the nucleic acids of the different phages revealed that the HindIII and AseI digests produced similar DNA fragment patterns. Additionally, SDS-PAGE analysis indicated that the isolated phages have similar structural proteins. The sequence BLAST results did not show any significant similarity with other previously identified phages. To the best of our knowledge, this study provides the first molecular characterization of L. garvieae phages.

Here, we report the first genome sequence of a new virulent phage of Microbacterium oxydans, termed vB_MoxS-ISF9, which was isolated from sewage. Transmission electron microscopy showed that the isolated phage, which has a hexagonal head of about 80 nm in diameter and a long non-contractile tail of about 240 nm, belongs to the family Siphoviridae. The vB_MoxS-ISF9 DNA was completely sequenced and found to be 59,254 bp in length, with a G+C content of 62.76% and 120 putative open reading frames (ORFs). The predicted protein products of the ORFs were identified, and their sequences were analyzed. In a comparison with all available phage genomes, vB_MoxS-ISF9 did not show any significant similarity to other previously reported bacteriophages. To the beast of our knowledge, this is the first report of the isolation and complete genomic sequencing of a virulent phage against a member of the genus Microbacterium.

Shigella spp. are water-borne pathogens responsible for mild to severe cases bacilli dysentery all around the world known as Shigellosis. The progressively increasing of antibiotic resistance among Shigella calls for developing and establishing novel alternative therapeutic methods. The present study aimed to evaluate a novel phage cocktail of lytic phages against extended spectrum beta lactamase isolates of Shigella species in an aquatic environment. The phage cocktail containing six novel Shigella specific phages showed a broad host spectrum. The cocktail was very stable in aquatic environment. The cocktail resulted in about 99% decrease in the bacterial counts in the contaminated water by several species and strains of Shigella such as Shigella sonnei, Shigella flexneri and Shigella dysenteriae. Achieving such a high efficiency in this in-vitro study demonstrates a high potential for in-vivo and in-situ application of this phage cocktail as a bio-controlling agent against Shigella spp. contamination and infections. [ABSTRACT FROM AUTHOR]