Advances in Applied Microbiology, Vol. 19

Advances in Applied Microbiology

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Wysocki P. Kowalczyk J. Labella J. Borrego D. Association of Myroides odoratimimus in immunocompromized piglets with post weaning multisystemic wasting syndrome Mamta Choudhary Binod Kumar Choudhary B. Bera S. Chaudhari D. Giri R. Ghosh S. Population structure and genetic diversity of Setosphaeria turcica from corn in Heilongjiang province, China Y. Jiang Q. Zhang E. Ali P. Pan L. They must be preserved by maintenance on agar slants stored in the cold or under sterilized mineral oil. These preparations require periodic transfer to fresh agar media. Possibilities for contamination or death are greater, and the cultures require more attention and concern on the part of curators.

These cultures must be transferred periodically to fresh agar slants and overlaid with sterilized mineral oil. Care must be taken in the overlaying technique so that every portion of the agar and culture is covered with the oil. If this is not done, dehydration of the medium occurs and the culture dies or becomes very difficult to retrieve.

The transferring process with oil-overlaid cultures is messy. Although we maintain only a small number of cultures in this fashion, including some patent cultures, they do constitute a problem and require consideration of alternative preservation methods.

Applied Microbiology and Biotechnology

The surge of development in hydrocarbon microbiology could lead to further problems should one attempt to preserve strains by this method. Utilization of the oil overlay by particular microorganisms which attack hydrocarbons is a possibility, No definitive data indicating qualitative productivity loss of cultures maintained under oil is known to us, To provide requestors, new agar slants must be prepared and grown up from the oil cultures. These cultures are subject to all the problems concerned with routine transfer of cultures, including run-down or loss of productivity.

STORAGE Storage of agar slant cultures or other preparations of microorganisms in deep freeze is another method by which patent cultures could be preserved for long periods of time. We do not use this method, although it might be useful for cultures not amenable to lyophilization. As with agar slants and cultures overlaid with oil, transfer to fresh media would be necessary at appropriate intervals. We have used the method 16 T. Any soil cultures received in connection with patent deposits are retained in our separate collection of original deposition material stored in the cold.

This process, pioneered here by Drs. Wickerham and K. Raper, has been used for the great majority of cultures maintained at the ARS Culture Collection for many years Preparations made in still are viable. Details of the process using relatively simple and inexpensive equipment are given in Wickerham and Andreasen 48 and Haynes et al. We use bovine blood serum exclusively as a suspending agent.

Thirty preparations of each patent strain are made initially. Before that material is completely exhausted, additional preparations are made, generally starting with material from the existing lot. We have seen no reason to change the methods or the simple equipment originally devised for the purpose. Bacteriophages recently reported to be present in some commercially produced bovine serum 12 may present a potential problem.

Milk or any other natural material used as a suspending agent also may contain such phages depending upon how it is processed. Whether the presence of these phages might exert effects on microorganisms during the process of suspending cells, spores, conidia, or hyphae; during the lyophilization process itself; or during the period that these materials are stored in uacuo in the cold is unknown.

The possibility of transduction or transformation presents itself. Research in this area is needed, and a reexamination of nonnatural suspending agents would be desirable. Also, when considering patent depositions in some other collections, one must bear in mind that certain countries may have scruples against using animal serum or milk as suspending agents. We do not presently use this method of preservation for any of our cultures. However, the slow but continuing accession of nonlyophilizable microorganisms has prompted us to consider this method for future use.

We are aware of one report of qualitative changes in productivity of a strain possibly related to its preservation with a liquid nitrogen process 2 4. Understandably, there is more information on viability of cultures after extended storage with lyophilized preparations than with liquid nitrogen preservation. Routinely, a viability check is made on each lot of lyophilized preparations made by our curators and records are carefully kept.

Advances in Applied Microbiology, Volume 53

No lot is acceptable unless viability can be demonstrated shortly after the materials are prepared. On receipt of complaint we check another preparation from the same lot. Only rarely have we been unable to get the strain to grow. Thus, we attribute most reports of nonviability of our preparations to lack of experience in handling lyophilized preparations or failure to follow the instructions accompanying each shipment. The viability checks also provide some characterization information as indicated before.

Advances in Applied Microbial Biology (AAMB) | Source Journals

Permanent availability has caused some discussion and elicited questions from potential depositors. Projection of the continued availability of progeny of microorganism strains deposited in culture collections throughout the world is dependent on many factors.

From a reasonable and practical standpoint, the deposit of strains in the permanent collection of the ARS Culture Collection would constitute maintenance in perpetuity. Likewise, deposit of strains in some of the other major collections throughout the world would constitute maintenance in perpetuity. NO culture collection in the world, however, can absolutely guarantee such maintenance.

Should a reputable collection be disbanded or cease to operate, the responsible parties undoubtedly would take steps to assure continued maintenance of their holdings by passing them on to other 18 T. We have salvaged several important collections of microorganisms in this manner. Once a strain becomes available, other collections may receive it and accession it. Progeny of the original deposit are available from a number of collections for many years to come, and continued availability is assured.

TIER 1 ARTICLE TYPES

Thus, we have duplicate collections maintained in other parts of our building complex and selected materials in other strategic locations. Thankfully, we have not yet had to rely on these collections. Other major collections probably have taken similar steps to protect their holdings.

Although the development of the lyophile and liquid nitrogen preservation processes guarantees that certain microorganisms will be available for many years to come, adequate safeguards should be instituted if not already in use. Records Maintaining records on ARS Culture Collection holdings takes up considerable time and space, as it does with other collections. Often, we must refer to records and correspondence of some 30 years ago to answer various kinds of questions that arise.

Breadcrumb

A separate file is maintained for each patent strain so that we can readily retrieve information on these microorganisms. The files are carefully checked with each culture request to determine whether the strain is available at that time. The system has worked quite well so far.

Volume 109. Advances in Applied Microbiology

In retrospect, adoption of computerized record keeping some 20 or 30 years ago would have made all of the major collections more useful. There is a prodigious amount of information available from these collections, and curators provide much assistance to other researchers in providing such information.

Computerization of the information today would be a formidable and fulltime task but may become necessary. Availability and Distribution With regard to availability and distribution of patent cultures, one wonders why such cultures are requested. AS touched on before, a prime issue with regard to patent cultures is their availability before and during application procedures and after granting of the patent. Distribution of these cultures is contingent on their availability. There is little question in our minds that precisely designated strains should be deposited in reputable culture collections and that they should be available to bona fide requestors when the U.

Therefore, before distributing patent cultures, careful checks of the files must be made. We would be hard put to distribute some cultures shortly after their receipt. Some culture preparations might take several weeks before any shipments could be made. Hayhurst 1 6 , in a discussion on legal aspects of industrial microorganisms in the British Commonwealth, has pointed out the desirability of depositing certain strains at the time of filing applications with no restrictions on availability save normal terms as to price, safety precautions, or the like.

These limitations, in addition to those cited above, all affect the time at which a culture could be shipped. Moreover, he indicates that not only the type strain but also the best strain or strains known to the inventor should be deposited. The Future A.

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