In case of bacteria like Escherichia coli the doubling time for it is 30min and within 6 hours of incubation log phase culture can be obtained, but in case of Xanthomonas oryzae it is 2 hours and its takes a longer period for it to reach the log phase, which is required criteria for phage infection. This makes it difficult for me to carry out the work with the isolation of bacteriophages. Can anyone suggest me how to overcome this issue?
How are you growing the strain now? Some basic strategies would be to try increasing the incubation temperature. For instance if you are growing it at 25 deg. C, try 28 and 30 deg. to see if it grows faster. Xanthomonas oryzae is an obligate aerobe, so good aeration is key. So it should be grown in a shaking incubator in an Erlenmeyer flask at least 10X larger than the culture volume to give a good surface to volume ratio. What media are you using? Some people use Wakimoto's medium X. oryzae, which is vitamin free, so you might try spiking it with say 1g/L of yeast extract to provide vitamins and other growth factors. You can also try "pre-conditioning" your medium to see if you can reduct the lag phase. One way to do this is to filter sterilize a late log phase culture of your X. oryzae and add a fraction of this to your new media to provide any trace nutrients that X. oryzae may require for rapid growth. Or you can so the same thing with an early log phase culture and once it is sterilized, see if you have a reduced lag phase if you re-innoculate the filter sterilized medium.
Good luck
You can try out following things: Check which one suits best
1. Use half strength, and quarter strength medium for culturing X oryzae.
2. Reduce amount of Carbon and Nitrogen sources of the medium.
3. Use complex sugar and avoid simple sugar in the media. You know, more complex carbon and nitrogen sources you will use the growth will slowed down.
4.Use minute amount of complex sugar as carbon source and more nitrogen source.
I'm sharing my experience while I used to work with actinomycetes fermentation. Hope it will work. Cheers!
How are you growing the strain now? Some basic strategies would be to try increasing the incubation temperature. For instance if you are growing it at 25 deg. C, try 28 and 30 deg. to see if it grows faster. Xanthomonas oryzae is an obligate aerobe, so good aeration is key. So it should be grown in a shaking incubator in an Erlenmeyer flask at least 10X larger than the culture volume to give a good surface to volume ratio. What media are you using? Some people use Wakimoto's medium X. oryzae, which is vitamin free, so you might try spiking it with say 1g/L of yeast extract to provide vitamins and other growth factors. You can also try "pre-conditioning" your medium to see if you can reduct the lag phase. One way to do this is to filter sterilize a late log phase culture of your X. oryzae and add a fraction of this to your new media to provide any trace nutrients that X. oryzae may require for rapid growth. Or you can so the same thing with an early log phase culture and once it is sterilized, see if you have a reduced lag phase if you re-innoculate the filter sterilized medium.
Good luck
I would go with John's suggestions. However, you should be aware that some species have laboratory doubling times which are species dependant and cannot be altered much by experimentation.
Respected John E Wertz
Thank you for your suggestion . I am using PSA Peptone sucrose agar and in that i add Sodium glutamate. Pre-conditioning the medium sounds good , i havent thought like that . But in a different way i thought of approaching it. The pathogen resides in the xylem vessels of the plant . so i thought something which mimics xylem sap will be a good nutritional medium. May be i should try pre-conditioning the medium as u have said.
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