Microbial strains: laboratory applications

Standardized microorganisms are widely used in microbiology and quality control laboratories across all industries. The many applications include, for example, fertility and selectivity tests on culture media, verification of the activity of bactericidal or bacteriostatic substances, the Microbial Limit Test on pharmaceutical products, verification of filter efficiency and control of identification systems bacterial.

International regulations give precise indications, specifying the strains to be used, and the methods for growth and propagation. An example is given by the European Pharmacopoeia, harmonized with the American one, which indicates a list of culture media for sterility checks and for microbiological product analyses, suggesting the strains for verifying fertility/selectivity:

• Aspergillus niger ATCC 16404

• Bacillus subtilis ATCC 6633

• Candida albicans ATCC 10231

• Clostridium sporogenes ATCC 11437 or ATCC 19404

• Escherichia coli ATCC 8739 • Pseudomonas aeruginosa ATCC 9027

• Salmonella enterica subsp. enterica serovar Typhimurium ATCC 14028 or Salmonella enterica subsp. enterica serovar Abony NCTC 6017

• Staphylococcus aureus ATCC 6538

Another far-reaching standard, ISO 11133:2014, dealing with the preparation of culture media for microbiology, also specifies a whole series of strains necessary for verifying the performance of the media themselves.

Collections

First of all, it should be remembered that the WDCM (World Data Center for Microorganisms) catalog exists which collects 194 strains from 49 collections, marking them in order to facilitate access to the reference microorganisms listed in the publication “Handbook of Culture Media for Food and Water Microbiology” (ISO TC 34 SC 9 Joint Working Group 5, Working Party on Culture Media of the International Committee on Food Microbiology and Hygiene (ICFMH-WPCM) In essence, the strains are assigned a WDCM number which is added, acting as a transversal link, to the identification codes that each strain has in the various reference collections.

Ready-to-use microorganisms or stock?

Laboratories have the possibility to choose whether to use: • internal stock libraries, generated from pure and standardized cultures, and maintained for subsequent routine use, or • microbial strains ready for use, freeze-dried and standardized, with the added possibility of being already titrated. In general, regardless of the type of strain chosen, some factors must be evaluated, such as the origin of the strain, conservation and the number of passages from the reference culture.

Passages

A “step” is represented by the transfer of the microorganism from a viable culture to fresh medium on which it can grow. Each subculture constitutes a step. The production of stock cultures from pure ATCC® culture represents the first step. Simply rehydrating a lyophilized strain or thawing a culture kept in the freezer is not a step, whereas it becomes one when the microorganism is reseeded in fresh medium. For the development of log libraries it is advisable to use logs with a low number of passages, the aim is to protect their characteristics. Typically, the specific rules for the various uses give indications regarding the number

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