A novel contaminant was isolated from Madin Darby Bovine Kidney (MDBK) cells. The organism was unable to grow on standard microbiological media by conventional techniques, but grew well in Dulbecco's Modified Eagle's Medium (DMEM) containing high glucose concentration. The organism formed a white biofilm on the bottom without any signs of turbidity. Upon genome sequence analysis of 16 S rDNA, the contaminant was identified as Sphingomonas sp. Shah, a member of the group α-Proteobacteria. Neutral red dye uptake method confirmed clear cytotoxic potential of the bacterium on A-549 cells. The organism was capable of invading and infecting different mammalian cell lines: MDBK, ZZ-R, 293-T, A549, and HeLa cells. Infected cells showed a variety of cytopathic effects including vacuolation at perinuclear area, cytoplasmic granulation and membrane blebbing. Microscopic analysis of the infected cells revealed the presence of cytoplasmic vacuoles harboring motile organisms. Apparently local serum preparations seem to be the source of this contamination, which is imperceptibly passed on from one culture passage to the other and ultimately leading to serious cytopathic manifestations.
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