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Method for Recovery of Viruses from Milk and Milk Products

National Center for Urban and Industrial Health, Cincinnati, Ohio

ABSTRACT

A modification of the plaque-forming unit assay was developed for recovery of viruses from milk and milk products. Viruses were assayed on primary cell cultures of Cercopithecus aethiops (African green) monkey kidneys under an overlay medium. The medium contained 0.95% clarified Ionagar no. 2, Eagle's minimum essential medium with nonessential amino acids in Hanks' balanced salt solution without phenol red, 2% fetal bovine serum, 0.19% NaHCO₃, 0.0015% neutral red dye, 0.51% MgCl₂ · 6 H₂O, and 1% sterile homogenized bovine milk. Geometric means and 95% confidence limits expressed as plaque-forming units/0.5 ml are presented for recoveries of Adenovirus 12, Herpes simplex virus, Influenza A virus, and New-castle disease virus from Eagle's medium and sterile milk. Recoveries of Adenovirus 12, Herpes simplex virus, Simian virus 40, and Reovirus 1 from milk, ice eream mix, chocolate milk, chocolate drink, or heavy cream were within ± threefold of viral plaque-forming units input, estimated by dilution. Elimination or reduction of bacterial and fungal contamination in the non-sterile milk or milk products was achieved by adding 1,000 units of Penicillin G per milliliter, 1,000 µg of streptomycin sulfate per milliliter, 50 µg of tetracycline hydrochloride per milliliter, and 0.5 µg of Amphotericin B per milliliter at least 48 hours before assay.