Procedure C.

1: Isolates from Procedure A will be subjected

to screening for anti-biofilm forming capacity against

Staphylococcus epidermidis, Candida albicans,

Escherichia coli, Aspergillus niger, Pseudomonas

aeruginosa and Chryseobacterium indologenes.

A loopful of LAB isolates will be inoculated into MRS Broth.

After 24 hours of incubation at 37 Degrees Celsius, bacterial

suspensions will be standardized to MacFarland 0.5 turbidity.

Then, 20 uL of every suspension will be loaded in the wells of

96-well plate, in triplicates each. The wells will be added with

180 uL of sterile MRS broth. The cultures will be incubated in an

anaerobic condition at 37 Degrees Celsius for 48 hours. After 48

hours of incubation, the cultures will be poured out and the

plates will be rinsed with sterile distilled water. The wells

will be added with 20 uL of 0.5 MacFarland standardized cultures

of S. epidermides, E. coli, P. aeruginosa, A. niger, C. albicans

and C. indologenes. The wells will be added with 180 uL of

sterile Nutrient broth. The set-up will be triplicated and will

be incubated for 48 hours at 37 Degrees Celsius. After 48 hours

of incubation wells will be rinsed with sterile distilled water

and will be added with Crystal Violet for 10 mins. After 10 mins,

the wells will be washed with sterile distilled water and will be
subjected to reading of absorbance using microplate reader at the

wavelength of 570 nm.