Bacteriophage Isolation

BELDOA, DE JESUS, EVIDENTE, KIM, REGAL

Rationale
Sewage water source of fecal coliforms and

bacteriophage Tryptic soy broth general purpose media, effective for E. coli Centrifuge remove particulate matter 0.22 M membrane filter to filter out bacteria

Rationale
Enrichment of sewage water
Often

the source material is inoculated directly into a broth medium which will encourage the proliferation of the desired organism increases the probability that colonies of the desired organism will be isolated upon subsequent streak-plating useful in recovering organisms which are in very low numbers

Rationale
Chloroform

and solvent Poorly soluble in aqueous media Frees bacteriophage stocks by releasing intracellularly accumulated cell wall hydrolase to dissolve membrane lipids of bacteria Soft NA jelly-like consistency; to visualize plaques produced after inoculation of sample on NA plates

Extractant

Results

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Theoretical results

Treatment Control (0.3 ml E. Coli) 1 (0.3 ml E.coli + 1 drop sewage water) 2 (0.3 ml E.coli + 5 drops sewage water)

Result + +++

What are the similarities in plaque technique and plate count in bacteria?
Plaque technique (used in

the experiment) Plaque: clearing zones created due to the replication of bacteriophage viruses

 Total

number of clearing zones = number of organisms per volume of liquid on the plate Similar to bacterial plate count : number of colonies instead of clearing zones

Cause of clearing zones: lysis of bacteria Bacteria: CFU (Colony forming units)

Bacteriophage: PFU (Plaque forming units)