Based on literature, I found more than a dozen of isolation media, but I cannot select which is (are) the best medium to have more chance for recovering maximum isolates.
The answer is very simple. You check on the environmental conditions (parameters) from which you wish to isolate the microorganism. Now develop a microcosmic environment in the laboratory from the 1oos of media that you have come across and see which is the one that resembles your microcosm. Now use that medium (a) with the incubation parameters to isolate the desired organisms.
you should first enrich the microbes in suitable media which contains organic and inorganic sources and concentration of the NaCl can be kept from 3-30%. This enrichment should continue for 7-10 days and then you should transfer them on the plate containing same composition. The bacteria who are moderately halophilic or extreme halophilic can survive during enrichment so they can only grow on the plate and incubate the plates for longer time. Extremophiles may take long time to grow.
Once colonies will be available, salt tolerance capacity can be analysed for growing them individually at high salt containing medium.
18-23 % Modified Growth Media, with peptone and yeast extract but without bile-salt, i think is mostly used one, at least in our lab. And, filtering the samples, then, enrichment of the sample before plating as mentioned by Shilpa Gupte is a good idea.
For isolation of Halophilic microorganisms the medium of choice can be the source from which you wish to isolate that microorganism. for example Sea water containing Glucose and Yeast extract etc.
I'm going to second Bina's suggestion of MGM (my lab has used it with great success) and her caution about bile-salts (if you get the wrong YE and peptone you will never get a halo to grow regardless of the media). I will also recommend that you download a copy of the Halohandbook (http://www.haloarchaea.com/resources/halohandbook/). It has some great media recipes and molecular biology protocols.
Hi Dr Coker,
Thanks for all your comments. I will have many things to explore and it is really great!
make extract by adding 50 grams of soil sample in 500 ml of sea water and boil for an hr then make up the volume to 1 ltr.
incubate your sample in that water as per your desired quantity for 48 hrs.
make nutrient agar plates using the sea water extract instead of distill water and add 20-30% of salt conc. and do spreading. you can get few isolates which can tolerate up to 40% salt or more.
Depending on the composition of the brine sample, you can select the medium.
If you are isolating halobacteria from marine salterns you can use the medium that is commonly used for H. salinarium or H. cutirubrum. The medium usually contains (g/1000ml): NaCl 250.0; MgSO4, 7H2O 2.0; KCl 0.5; CaCl2 0.02; Bacteriological peptone 10.0; yeast extract 0.5; glycine 0.2; pH 7.5. The incubation time is at least one week.
You may modify the medium by using different concentration of salts or addition of substrates such as starch.
Wish you best in your search for halophiles!!!
If you're having trouble getting initial growth, another strategy you might employ is to sterile filter your environmental waters and use that as a growth medium. Simply add an appropriate electron donor and inoculate with a small volume of your sample. This is advantageous if you're unsure of the precise optimal growth conditions as you'll have higher levels of viable cells to work with when later optimizing your growth media.
If you're looking for a specific media, give this one a try. I used it to isolate halophiles from Mono Lake (California):
https://www.researchgate.net/publication/283326936_Aliidiomarina_minuta_sp_nov_a_haloalkaliphilic_bacterium_that_forms_ultra-small_cells_under_non-optimal_conditions
Good luck!
Article Aliidiomarina minuta sp. nov., a haloalkaliphilic bacterium ...
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