Improved bactericidal efficacy and thermostability of Staphylococcus aureus-specific bacteriophage SA3821 by repeated sodium pyrophosphate challenges

As antibiotic resistance is being a threat to public health worldwide, bacteriophages are re-highlighted as alternative antimicrobials to fight with pathogens. Various wild-type phages isolated from diverse sources have been tested, but potential mutant phages generated by genome engineering or random mutagenesis are drawing increasing attention. Here, we applied a chelating agent, sodium pyrophosphate, to the staphylococcal temperate Siphoviridae phage SA3821 to introduce random mutations. Through 30 sequential sodium pyrophosphate challenges and random selections, the suspected mutant phage SA3821M was isolated. SA3821M maintained an intact virion morphology, but exhibited better bactericidal activity against its host Staphylococcous aureus CCARM 3821 for up to 17 h and thermostability than its parent, SA3821. Sodium pyrophosphate-mediated mutations in SA3821M were absent in lysogenic development genes but concentrated (83.9%) in genes related to the phage tail, particularly in the tail tape measure protein, indicating that changes in the tail module might have been responsible for the altered traits. This intentional random mutagenesis through controlled treatments with sodium pyrophosphate could be applied to other phages as a simple but potent method to improve their traits as alternative antimicrobials.

Characterization and Genome Sequence of a Genetically Unique Escherichia Bacteriophage vB_EcoM_IME392

In this study, a novel Escherichia coli-specific bacteriophage vB_EcoM_IME392 was isolated from chicken farm sewage in Qingdao, China. The genome of IME392 sequenced by Next Generation Sequencing is found to contain 116,460 base pairs in length with a G+C content of 45.4% (deposited in GenBank under the accession MH719082). BLASTN results reveal that only 2% of the genome sequence of IME392 shows homology to known phage sequences in GenBank, which indicates IME392 is a very novel bacteriophage. Transmission electron microscopy showed that IME392 belongs to the family Myoviridae. The host range, multiplicity of infection and one-step growth curve was also determined.

The morphological and biological characteristics of a virulent PI phage isolated from slaughterhouse sewage in Shiraz, Iran

Background and Objectives: Foodborne pathogens are among the serious problems all around the world and thus a novel and natural strategy to control and to inhibit such pathogens is highly demanded nowadays. The aim of this study was to iso- late a specific bacteriophage of Escherichia coli O157:H7 from sewage in Fars province, Iran to determine its morphological and antimicrobial activities. Materials and Methods: In order to isolate the bacteriophage of E. coli O157:H7, 10 samples of slaughterhouse wastewa- ters were used. Double-Layer Agar method was employed to isolate the bacteriophage. To identify the fine structure of the bacteriophage, electron microscope was employed. Host range and antibacterial activity of the phage was also investigated, in vitro. Results: The morphological and biological characteristics of a virulent Siphoviridae phage, PI, are reported. It was found that infection of E. coli O157:H7 strains with this specific bacteriophage produce clear plaques. In the one-step growth anal- ysis, it was confirmed that the phage has been characterized with a very short rise period (around 15 min), an average burst size of 193 PFU/cell, high infectivity and potent lytic action. The bacteriolytic activity of PI was also investigated, in vitro. It was also clarified that at the MOI of 100, 10 and 1, the phage rapidly lysed the bacterial cells within 0.5 or 2 h. Conclusion: These results indicate that the phage PI is a newly discovered phage against E. coli O157:H7 in Iran which may be recommended to use as bio-control purposes.

Complete Genome Sequence of the Streptomyces-Specific Bacteriophage BRock

ABSTRACT The complete genome sequence of the unique virulent bacteriophage BRock, isolated from compost on Streptomyces sp. strain SFB5A, was determined. BRock is a myovirus with a 112,523-bp genome containing a GC content of 52.3%. There were 188 protein-coding genes predicted, including structural and enzymatic proteins, but none predicted for lysogeny. Twenty-nine tRNAs were predicted.

Determination of Optimum Survivability Factors of Highly Pathogenic Vibrio cholerae 01 Serogroup-Specific Bacteriophage JSF4ϕ

Cholera is severe watery diarrhea caused by pathogenic V. cholerae 01 or 0139 serogroups. In each year, 2.9 million people are affected by cholera worldwide and 95000 deaths occur from the disease annually. In Bangladesh, around 100000 people are affected by this disease and approximately 4500 deaths occur each year. In this study, a novel V. cholerae 01 serogroup-specific bacteriophage JSF4ϕ was used. This phage was able to lyse both the clinical and environmental pathogenic V. cholerae 01 serogroup strains and one of our previous studies demonstrated that the seasonal outbreaks of cholera caused by V. cholerae 01 serogroup strains in Bangladesh are mostly regulated by this bacteriophage. In this current study, we determined the optimum survivability factors of JSF4ϕ bacteriophages. This study showed that the temperature 2500C, pH 7 and normal saline are the optimal survivability factors for JSF4ϕ bacteriophages because, at these conditions, we have got the maximum number of plaque-forming units (PFU/mL) of these bacteriophages. This study also showed that the JSF4ϕ bacteriophages can survive at a wide range of temperature, pH and salinity. So, the study presented here may have an impact on the controlling of cholera epidemics caused by environmental and clinical pathogenic V. cholerae 01 serogroup strains if we can use JSF4ϕ bacteriophages as a biocontrol agent. This study may also have profound implications for future studies of JSF4ϕ bacteriophages as a good food additive or in phage therapy for its efficient lysing capacity against the pathogenic V. cholerae 01 serogroup strains.

Methods for Extraction and Detection of Bacteriophage DNA from the Sputum of Patients with Cystic Fibrosis

There is increasing interest in the pulmonary microbiome’s bacterial and viral communities, particularly in the context of chronic airway infections in cystic fibrosis (CF). However, the isolation of microbial DNA from the sputum from patients with CF is technically challenging and the optimal protocols for the analysis of viral species, including bacteriophage, from clinical samples remains challenging. Here, we evaluate a set of methods developed for processing and analyzing sputum from patients with CF with a particular emphasis on detecting bacteriophage viron-derived nucleic acid. We evaluate the impact of bead-beating, deoxyribonuclease digestion, and heating steps in these protocols focusing on the quantitative assessment of Pseudomonas aeruginosa and Pf bacteriophage in sputum as a proof of concept. Based on these comparative data, we describe an optimized protocol for processing sputum from patients with CF and isolating DNA for PCR or sequencing-based studies. These studies will facilitate future assessments of bacteriophage and bacteria in sputum from patients with CF.Author SummaryPatients with cystic fibrosis (CF) have thickened secretions and develop chronic infections in their airways. While we culture bacterial pathogens from expectorated sputum this method favors detection of certain organisms. There is greater understanding that the microbiome with in the airway of patients with cystic fibrosis is varied and contains more than just the bacteria that we selectively culture. Processing sputum is difficult as it is very thick. There are many different ways to process sputum depending on what aspect of the sputum is to be studied. We have an interest in a specific bacteriophage, or a virus that infects bacteria. We sought to find the best method to take sputum from a CF patient and extract DNA so that we could detect both bacteria and bacteriophage DNA. We compared different methods that included different combinations of heat, mechanical homogenization, chemical lysis and DNA extraction by commercially available kits. We describe the method we found easiest to execute and produced the best yield for detection of both bacteria and bacteriophage. Our purpose of publishing this method in detail is to facilitate further study of viral and bacterial communities in the sputum of patients with CF.

New method of evaluating specific bacteriophage activity using HLE-3 cell culture

In the presented study, we evaluated the efficacy of specific bacteriophage activity in vivo for resolution of infections caused by Staphylococcus aureus, Pseudomonas aeruginosa and Acinetobacter baumanii. Pseudomonas aeruginosa, Acinetobacter baumannii. It was demonstrated that administration of a specific bacteriophage with high lytic activity in an hour leads to a decrease in the adhesive activity of the microorganisms. The duration of the experiment was one hour.