Minimum essential medium (mem)

Enrichment of milk samples was performed in Luria Bertani (LB) broth as described earlier [18 (link)]. 100 μl of the enriched sample was spread onto modified Edwards medium (MEM) (Himedia, India) and incubated aerobically overnight at 37°C. Black color colonies characteristic of Enterococcus spp. obtained on MEM were screened for bacterial morphology by Gram's staining. At least three colonies showing characteristics of Enterococcus spp. were purified by subsequent streaking onto MEM. Crude genomic DNA was extracted from the purified colonies by boiling method and subjected to PCR targeting ddl gene [19 (link)] for the identification of E. faecalis using the primers provided in Supplementary Table S1. PCR reaction was adjusted to 20 μl volume with 10 μl 2X GoTaq® G2 Green Master Mix (Promega, USA), 10 pmol of each primer (Supplementary Table S1), and 2 μl of DNA templates. PCR was conducted in an ASTEC 482 thermal cycler (Japan) with an initial denaturation at 95°C for 5 min followed by 30 cycles of denaturation at 95°C for 30 sec, annealing at 54°C for 30 sec, extension at 72°C for 1 min, and a final extension step at 72°C for 5 min. Representative isolates which were positive for the ddl gene were further confirmed by sequencing of 16S rRNA using the primers 8F and 1492R [20 ] (Supplementary Table S1).

AZT was obtained as a gift sample from Cipla Ltd. (Goa, India). Cholesterol (CHO), fluorescein isothiocyanate (Isomer I) (FITC), SL, fetal bovine serum (FBS) and minimum essential medium Eagle (MEM) were procured from HiMedia Laboratories Pvt. Ltd. (Mumbai, India). SA, OA, and PA were purchased from Sigma-Aldrich (Bangalore, India) and butylated hydroxytoluene (BHT) was obtained from Qualigens Fine Chemicals (Mumbai, India). U-87 MG cells were procured from National Center for Cell Science (Pune, India). All other chemicals used were of analytical grade.