I regularly have to enumerate bacteria (E. coli and pathogens) from soil and sediment samples. However, I still do not have conclusion for suitable practice. Normally, the enumeration would first making serial dilutions of samples, then inoculate to the enrichment media, and incubate in shaking incubators or media plates.

I found out that for E. coli, the soil extaction protocol of using slow speed (160 rpm) shaking by Kingsley and Bohlool (1981) was more suitable than using vortex. With the maximum speed of vortex in serial dilution step, there was no E. coli growth on plates, and I guess that the particles in samples might cause cell damages.

Then, I feel insecure when I decide to enumerate bacteria by directly put soil and sediments into enrichment media and incubate in shaking conditions for overnight or more. I always end up using low speed (160 rpm) than 250 rpm that I generally use for cell cultivation. Also, when I take the enriched culture to inoculate the next media, I am not sure that should I mix the enriched culture that has soil/sediment particles with vortex or not, and end up just shake by hands.

Anyone has suggestions on this?

Thank you so much

Ref:Kingsley and Bohlool (1981), Release of Rhizobium spp. from tropical soils and recovery for immunofluorescence enumeration