I presume that you have tested that the actual strain you are using is motile? But you have all the parts to troubleshoot this. Trade plates, strains etc with your lab mates and see if their strains work in your hands, and vice verse. This should be simple to isolate the problem given that others can get it to work in your lab.

Hello Michael

I did the things you recommend like trading plates, asking my lambastes to prepare media vs me preparing my media to check if handling issues are there, we tried to inoculate both mine and their perfectly working strains to check if there is an issue with the bacteria. The only thing I did not do was specifically testing motility using motility tests. I though the bacteria all of a sudden won't lose its motility. But that is something I plan to do in fact today itself. I will let you know how it goes.

Thank you very much for your advice. It really helped me understand that I am still on the right path to troubleshoot it :)

At least E. coli will often lose its motility quite easily.

I will join @Michael answer, take a working strain from a college and see if it works in your hands

Hello

I was able to test the motility of my culture, as specified by Michael and determined that my culture infact is very well motile. I have requested my lab mate to prepare the entire experiment with his handling and inoculate mine and his culture. This will suggest if there is issue with my handling or not.

Thank you so much for both your insights, I think I am closer to identify the issue now.

:)