My Cart
Your Shopping Cart is currently empty. Use Quick Order or Search to quickly add items to your order!
Living Organism Care Guide: Halobacterium
Living Care Information
Halobacterium salinarum
commonly known as Halophile,
extremophile
Quick Start Information
Open the shipping container and locate your cultures. Check tube cultures, ensuring that the cap is tightly in place. Check plate cultures, ensuring that the plate is unbroken and securely taped.
Check for any damage on the bag holding samples of brine inclusion crystals.
Halobacterium cultures can be held at room temperature for several weeks before subculturing is required. For longer term storage, seal and store in a refrigerator (4° C).
The speed at which subcultures develop is dependent on temperature. Incubating at 42° C produces subcultures in approximately 2 weeks.
About the Organism
- Despite its name, Halobacterium is not classified as a bacterium, but a member of the domain Archaea.
- Halobacterium is an extremophile not only because of its existence in very salty environments but also because of its extremely high tolerance to UV exposure.
- Halobacterium is naturally found in hypersaline lakes and areas used for making salt,called salterns. As evaporation progresses, the water in salterns begin to turn purple or pinkish in color due to the presence of high levels of halophilic Archaea, including Halobacterium species. They have also been isolated from ancient salt, including from Permian Period halite (250 million years old).
- Domain: Archaea
- Phylum: Euryarchaeota
- Class: Halobacteriales
- Family: Halobacteriaceae
- Genus: Halobacterium
- Species: sp.
- Strain: NRC-1
Preparation
Halobacterium cultures will remain viable for 3 to 4 weeks at room temperature. Be sure to have media and basic microbiological supplies available if you plan to start your work immediately after receipt.
Housing
Halobacterium can be cultivated successfully on plates, standard slants, or in broth. We recommend Halobacterium agar or Halobacterium broth. The amount of agar provided is sufficient to pour 5 plates or 15 to 20 tubes. Broth tubes contain 5 mL. Plan to use media within 3 months of receipt for best results. Please note that the broth contains salt to the point of saturation. Some precipitate is expected and does not indicate media contamination.
When pouring plates using Halobacterium agar, use eye protection, a protective lab coat, and heat-resistant gloves. See our video “How to Melt and Pour Agar Plates” for a demonstration.
- Loosen the cap on a bottle of agar and set the bottle in a pot of room temperature water on a heat source. Make sure the water level is even with or slightly above the level of the medium in the bottle. Bring the water to a boil. Leave the bottle in the boiling water until the agar completely melts (about 30 minutes).
- Allow the bottle to cool to 55° C; it should feel comfortably hot to the touch.
- Disinfect the work surface with a disinfectant such as bleach or alcohol. Wash hands thoroughly. Align the sterile plates along the edge of the clean work surface.
- Remove the cap and flame the mouth of the bottle of agar. While pouring the plates, lift the lid of each petri dish just enough to pour in the molten agar. Pour just enough to cover the bottom of the plate. After pouring each plate, immediately replace the lid to prevent contamination. One bottle should pour 4 to 5 plates.
- Leave the plates undisturbed until they solidify (about 1 hour).
- 6. Store plates in an airtight container in a refrigerator and use within 4 weeks of pouring. When preparing to streak the plates, let the plates warm up to room temperature and make sure there is no condensation on the lid. The water will lyse cells. Note: Always store plates in airtight containers—even during incubation. Otherwise, they dry out and salt crystals form.
Feeding
If you are planning to culture large quantities of Halobacterium in broth with shaking, you will lose volume due to evaporation. Replace the volume with sterile broth.
Maintaining and culturing
Streak the plates or spread the surface of the plates with liquid from a broth culture. Once inoculated, incubate at either 42° C (preferred) or 37° C. At 42° C, cultures are ready in 7 days. At 37° C, they take about 1 to 2 weeks. If the plates incubate at room temperature, results may take 2 weeks or more, depending upon the temperature of the classroom.
Disposal
When you are finished with your materials, please dispose of them in one of the following ways:
- Soak materials in a 20% bleach solution for 10 minutes.
- Place the materials in a 70% ethanol bath for 24 hours.
- Autoclave the materials at 121° C for 15 minutes in an autoclavable bag. Petri dishes will melt in the autoclave, so be sure to contain materials and close securely before autoclaving.
Biosafety
According to the U.S. Public Health Service’s definition of Biosafety Level 1, Halobacterium cultures are considered to be non-pathogenic. They would not be expected to cause harm to healthy people or to the environment.
Even though considered safe and non-pathogenic, any microorganism should be always treated as if it were a potential pathogen. Laboratory work should be done only in accordance with prudent laboratory safety precautions, standard microbiological practice, and aseptic technique, and under the supervision of a person familiar with such precautions and practices. Gloves should be worn, work surfaces should be disinfected before and after use, and hands should be washed thoroughly after glove removal. When finished, all materials should be properly disinfected and disposed of safely.
Video
FAQs
How can I prepare a Halobacterium culture for Gram staining?
Halobacterium NRC-1 is reported as a gram-negative organism and has no bacterial peptidoglycan.
If samples need to be prepared for Gram stain, we advise the following:
- Haloarchaens require >2M NaCl for membrane stability. Thus, there is a possibility for lysis during the gram staining sequence if samples are not completely heat fixed.
- Cells naturally exhibit a pink color, so staining components may not be apparent after gram staining is complete.
- Halobacterium media contains high levels of salt that may affect the efficacy of stain mixes.
The cultures I prepared from the brine inclusion crystals are not growing.
The brine crystals can contain vastly different amounts of inoculum, so cultures started from them must be inoculated heavily. Our recommendation is 2 to 3 crystals per 5 mL of Halobacterium broth. Beyond that, Halobacterium takes a long time to grow.
The fastest growth (in our hands) happens at 42° C in flasks with shaking. Shaking promotes growth, but also promotes evaporation. We find that it is necessary to aseptically replace some of the media periodically if using a shaking incubator for Halobacterium cultivation. Once the culture begins to turn pink, it has reached a high enough inoculum that it can be used for experiments or to make additional subcultures.
In what way is Halobacterium an extremophile?
Extremophiles flourish under conditions that would kill most other organisms. For example, one type of extremophile—thermophiles—thrive in the geothermal springs in Yellowstone National Park at temperatures above 80° C. Halobacterium is a halophile (salt lover). It grows optimally from 3.4M to 5.1M (20% to 30%) NaCl. Most organisms grow optimally at less than 0.2M NaCl.
Can I culture Halobacterium on nutrient agar?
Halobacterium only grows on a hypersaline medium. It cannot grow on standard bacteriological media. Few other organisms can grow on media formulated for Halobacterium.
Need help?
We want you to have a good experience. Orders and replacements: 800.334.5551, then select Customer Service. Technical support and questions: caresheets@carolina.com
