Propagating the missing bacteriophages: a large bacteriophage in a new class

Area of Science:

• Microbiology
• Virology
• Genomics

Background:

• The isolation of soil-borne bacteriophages is limited compared to microscopic observations, a phenomenon known as the great plaque count anomaly.
• Conventional microbiological techniques often fail to detect certain bacteriophages due to their unique properties.

Purpose of the Study:

• To resolve the great plaque count anomaly by developing a novel isolation method.
• To isolate and characterize a previously undetectable soil-borne bacteriophage.

Main Methods:

• Utilized propagation in ultra-dilute agarose gels for bacteriophage isolation.
• Characterized the isolated bacteriophage, 0305phi8-36, including its morphology and genome size.
• Assessed plaque formation dependence on agarose gel concentration and lysis in liquid cultures.

Main Results:

• Successfully isolated a novel Bacillus thuringiensis bacteriophage, 0305phi8-36, with distinct morphological features (large head, tail, tail fibers) and a 221 Kb genome.
• The bacteriophage exhibited high dependence of plaque size on agarose gel concentration and did not propagate in traditional gels or cause visible lysis in liquid cultures.
• 0305phi8-36 aggregates and is likely invisible to standard filtration or centrifugal isolation methods.

Conclusions:

• The novel isolation method using ultra-dilute agarose gels effectively addresses the great plaque count anomaly.
• Bacteriophage 0305phi8-36 represents a new genomic class, expanding knowledge of environmental virus diversity.
• This discovery has implications for future exploration of microbial virus diversity.