Background

Discovery of Novel Phages

Phages are incredibly diverse, and many remain undiscovered. Natural environments such as soil, water, and clinical settings harbor a vast array of phages with unique characteristics. Phage screening allows researchers to explore these environments and discover novel phages with potential therapeutic or biotechnological applications. For example, phages isolated from soil or wastewater may have unique lytic mechanisms or host ranges that could be exploited to treat antibiotic-resistant infections or for bioremediation processes.

Characterization of Lytic Profiles

This involves determining whether a phage has broad-spectrum or narrow-spectrum lytic activity. Broad-spectrum phages can infect and lyse multiple bacterial species or strains, making them useful for treating mixed infections or when the specific pathogen is unknown. In contrast, narrow-spectrum phages target specific bacterial strains with high precision, which is advantageous for treating infections caused by a single, well-defined pathogen. Understanding the lytic profile of a phage is essential to optimize its use in phage therapy or other applications.

Safety and Efficacy Assessment

Before phages can be used in clinical or industrial settings, their safety and efficacy must be thoroughly evaluated. Phage screening is the first step in this process, which facilitates the identification of phages that are not only effective against the target bacteria but also safe for human or environmental use. Screening helps to eliminate phages that may carry harmful genes or that could potentially cause adverse effects when used in therapeutic or biotechnological contexts.

Sources of Phages

• Natural Environments: Phages are isolated from diverse natural habitats rich in bacterial hosts, including soil, freshwater, marine environments, and wastewater.
• Clinical Settings: Targeted phage isolation can be performed from clinical samples such as human and animal feces, saliva, dental plaque, wound exudates, as well as skin and mucosal surfaces.

Figure 1. Phage distribution and abundance in three ecosystems. (Dion et al., 2020)

Phage Isolation Methods

Phage isolation from these sources involves several steps, including sample collection, enrichment, purification, and characterization. Common methods include:

• Direct Plating: Applying samples directly to bacterial lawns to detect plaques, ideal for high-phage samples like stool but may miss non-plaque-forming phages.
• Enrichment Cultures: Incubating samples with specific bacteria to amplify low-abundance phages, often requiring multiple rounds for sufficient titers.
• Plaque Assay: The standard method for detecting and quantifying phages by mixing filtrates with bacteria and spotting clear plaques on agar plates, indicating active phages and their concentration.

Figure 2. Flow chart illustrating the basic techniques for phage isolation. (Van Charante et al., 2019)

Creative BioMart Microbe offers natural phage discovery and isolation services. Our expert team specializes in isolating wild-type phage from a variety of environments—tailored to your target bacteria, application needs, and project goals. Contact us today to discuss your specific needs and how our services can accelerate your research efforts.

Service Procedure

Service Details

• Sample Collection & Processing: Systematic collection from environments rich in bacterial hosts such as soil, water, wastewater, and clinical specimens (e.g., feces, saliva, wound swabs).
• Phage Enrichment: Selective amplification of phages using targeted bacterial hosts to increase phage abundance when naturally low.
• Direct Plating & Screening: Rapid detection of lytic phages by applying samples directly to bacterial lawns and identifying plaques.
• Plaque Purification: Isolation of individual phage clones through serial plaque picking for purity and consistency.
• Phage Characterization: Initial phenotypic and genetic analysis to confirm host range, morphology, and stability.
• Custom Isolation Strategies: Tailored approaches based on project goals, including multi-host screening and selective enrichment.

Our Advantages

• Expertise: Our team consists of experienced microbiologists and molecular biologists with extensive experience in phage research.
• Customization: We tailor our services to meet your specific research objectives, whether it's targeting a particular pathogen or constructing a specialized library.
• Innovation: We employ cutting-edge technologies and methodologies to remain at the forefront of phage research.
• Quality Assurance: Rigorous quality control measures guarantee the reliability and reproducibility of results.
• Compliance: All procedures adhere to relevant regulatory standards, ensuring ethical and responsible research practices.
• Comprehensive Support: From project initiation to data interpretation, we provide end-to-end support to facilitate your research journey.
• Global Reach: Partners with academic, biotech, and pharmaceutical customers on every continent to deliver consistently high quality, customized phage solutions with global scalability and support.

Case Study

Case Study 1: Polyvalent Myovirus (vB_STM-2) for Salmonella typhimurium biofilm removal.

A potent lytic phage, vB_STM-2, was isolated for the control of multidrug-resistant Salmonella typhimurium. It exhibited a broad host range, strong biofilm reduction (up to 93.4%), and stability at various temperatures and pH levels. vB_STM-2 significantly reduced Salmonella on chicken meat during both short- and long-term storage, making it a strong candidate for antimicrobial and food safety applications.

Figure 3. Salmonella typhimurium phage characterization. (A) Clear, large circular plaques (4.5 mm in diameter) were produced on the double-layered agar plate. Electron micrographs at 80,000× of (B) vB_STS-1 phage (Siphovirus), (C) vB_STM-2 phage (Myovirus) and (D) vB_STS-3 phage (Siphovirus). (Abdelhadi et al., 2021)

Case Study 2: Phage cocktails against Paenibacillus larvae for American foulbrood control.

Using a community science approach, researchers in New Zealand have isolated eight Paenibacillus larvae strains and 26 novel phages native to the region to combat American foulbrood (AFB), a deadly honey bee disease. With the use of antibiotics banned in many areas, including New Zealand, these phages provide a natural alternative. Genomic analysis and host range testing led to the formulation of effective phage cocktails that show great potential to protect honey bees from AFB in a safe, targeted and antibiotic-free manner.

Table 1. Details of 26 Paenibacillus larvae phages discovered, sequenced and annotated. (Kok et al., 2023)

In this study, the phage plaque was purified, phage lysates were prepared, and phage DNA was extracted and sequenced. The phage detection workflow is shown in Figure 4.

Figure 4. Phage discovery (A) Schematic of phage enrichment and isolation process; (B) Positive spot tests after enrichment; (C) Representative TEM image of Phage Lilo (Harrison cluster); (D) Representative TEM image of Phage Ollie (Vegas cluster). (Kok et al., 2023)

FAQs

References:

1. Abdelhadi IMA, Sofy AR, Hmed AA, Refaey EE, Soweha HE, Abbas MA. Discovery of polyvalent Myovirus (vB_STM-2) phage as a natural antimicrobial system to lysis and biofilm removal of Salmonella typhimurium isolates from various food sources. Sustainability. 2021;13(21):11602. doi:10.3390/su132111602
2. Dion MB, Oechslin F, Moineau S. Phage diversity, genomics and phylogeny. Nat Rev Microbiol. 2020;18(3):125-138. doi:10.1038/s41579-019-0311-5
3. Kok DN, Zhou D, Tsourkas PK, Hendrickson HL. Paenibacillus larvae and their phages; a community science approach to discovery and initial testing of prophylactic phage cocktails against American Foulbrood in New Zealand. MRR. 2023;2(4):N/A-N/A. doi:10.20517/mrr.2023.16
4. Van Charante F, Holtappels D, Blasdel B, Burrowes B. Isolation of bacteriophages. In: Harper DR, Abedon ST, Burrowes BH, McConville ML, eds. Bacteriophages. Springer International Publishing; 2019:1-32. doi:10.1007/978-3-319-40598-8_14-1