Rehydration and Growth of freeze dried cultures
Each yeast is dispatched as a small quantity of freeze-dried culture sealed under vacuum in glass ampoules. Ampoules should be opened in the following way:
- 1). Check the number on the label inside the ampoule.
- 2). Score the glass with a suitable file or scoring impliment at the level of the cotton wool plug.
- 3). Wipe the ampoule with alcohol.
- 4). Apply the end of a red-hot glass rod to the file mark to crack the glass. Leave the glass rod in contact with the ampoule for 2-3 seconds if the ampoule does not crack immediately. If the ampoule still fails to crack repeat the process until successful.
- 5). Remove the tip of the ampoule and cotton wool plug and place in a container for subsequent sterilization before disposal.
- 6). Using a sterile Pasteur pipette, add about 0.5ml YM broth (Difco 0711-01) or malt extract to the dried material.
- 7). Gently resuspend the dried culture and transfer to a culture bottle containing approximately 10ml of the same medium. In order to prevent pH shock etc. it is recommended that no more than 10mls nutrient broth be used for initial growth of the culture.
- 8). Unless otherwise indicated, most yeasts should be incubated at 25°C. Growth may be slow immediately after resuscitation. It may be necessary to incubate cultures for at least 5 days before discarding. Growth is usually stimulated by aeration which may be achieved by shaking or rocking the culture.
- 9). The discarded ampoule should be placed in a container for subsequent sterilization before disposal.
All NCYC strains (except where indicated) can be routinely grown in YM broth (Difco 0711-01). Other suitable nutrient media include:
- YM: 0.3% yeast extract, 0.3% malt extract, 0.5% peptone, 1% glucose
- YEP-glucose: 0.5% yeast extract, 0.5% peptone, 1% glucose
- Malt Extract: 0.3% malt extract, 0.5% peptone
- Sabouraud’s Glucose: 1% peptone, 4% glucose
- YPD medium: 1% yeast extract, 2% peptone, 2% glucose
- Yeast Nitrogen Base (Difco 0392-15-9): a chemically defined medium to which a carbon source must be added.
- Yeast Nitrogen Base without amino acids (Difco 0919-15-3), which can be supplemented with appropriate amino acids or other source of nitrogen (useful for some genetically defined strains). A carbon source must be added.
Agar when required is added to a final concentration of 1.5 – 2.0% (All percentages are given weight/volume)
More information about these media is given by J P van der Walt & D Yarrow, ‘Methods for the isolation, maintenance, classification and identification of yeasts’ in ‘The Yeasts . A Taxonomic Study’, third edition by ed. N J W Kreger-van Rij, Elsevier, (1984) pp 45-104, and in ‘Yeasts: Characteristics and Identification’, third edition by J A Barnett, R W Payne & D Yarrow, Cambridge University Press, (2000).
Unfortunately most of the brewing yeast have been received on a confidential basis and the depositors or customers who use the strains do not usually give us much feedback on how the yeast worsk as regards giving the final product its characteristics. We are therefore unable to give customers details on exactly which strains in the NCYC have been used to brew particular beers.
It is however usually possible to get a yeast from the collection which matches the charateristics of an older strain using the ‘Tall Tube Codings’ which are given with most of the strains. Our customers usually use these to select a yeast which they think is suitable for their purposes. It is difficult to predict exactly how a yeast will behave and what the character of the final brew will be, since this tends to vary depending on the other ingredients used and the exact brewing process. We usually suggest that a customer picks one or two yeast which look suitable and try trial brews. The full explanation of the ‘Tall Tube Codings’ can be found in the brewing yeast section of the web site.
Pathogenic yeasts and fungi from clinical sources are held at the National Collection of Pathogenic Fungi. In general the NCYC holds non-pathogenic yeasts but there are certain exceptions to this and the following comments are included as a guide to users. It must be noted that pathogenicity is not an absolutely defined characteristic and so the following should be used as a guide rather than a definitive account of the subject.
Many kinds of microorganism may become opportunistic pathogens if they gain access to the human bloodstream. If the subject has an impaired immune system the consequences of such invasion are very serious and may be fatal.
The most pathogenic yeasts are certain Candida species and Filobasidiella neoformans, the asexual state of which is called Cryptococcus neoformans. The latter species is not held by NCYC.
Hurley et al (1987) list the pathogenic yeasts of candidosis in probable descending order of virulence for man as: C.albicans, C.tropicalis, C.stellatoidea, C.glabrata, C.krusei, C.parapsilosis, C.guilliermondii, C.viswanathii, Clavispora lusitaniae (Candida lusitaniae) and Rhodotorula mucilaginosa (Rh.rubra). (Ref: ‘The Yeasts’, Vol 1. ed. A H Rose and J S Harrison. Academic Press, 1987: Chapter 4). However only a few Candida species are markedly pathogenic. Some species of Malassezia, Trichosporon and Geotrichum may also be pathogenic.
A number of other yeast species not mentioned here have also been described as pathogens under particular circumstances and it is advised that a literature search is made if a decision is required on any particular strain or species.
Customers should always employ good microbiological practice when working with yeast cultures and use ‘Category 2’ containment when working with the Candida strains listed above, or any yeast strain suspected of being a pathogen. Full details can be found in: ‘Categorisation of biological agents according to hazard and categories of containment’, Advisory Committee on Dangerous Pathogens, HSE Publications, ISBN 0 7176 1038 1. The NCYC will be pleased to offer advice on suitable containment levels.
Please note that the NCYC is prohibited from sending cultures which are regarded as human or animal pathogens, or that are classified as ‘dual use’ to certain countries.
The NCYC is frequently asked for information on a culture we have which is known as ‘Bees Wine’. This was a fermented drink which was most often produced in home kitchens and was probably most popular in the 1920’s to 1950’s. The culture was usually kept in a glass container by a window and grown in a mixture of water, brown sugar and black treacle (there are several variations on the exact ingredients used). It was usually drained once a week and fresh water and sugar were added. As the culture naturally multiplied any excess was either discarded or passed on to others to begin new ‘Bees wine’.
According to our old records ‘Bees Wine’ is a mixture of yeasts and bacteria. The bacteria are Lactobacilli and an unknown Gram positive rod that forms a gelatinous sheath that coils and traps the other cells in it.
This is also responsible for a thick ‘scum’ which forms on the surface of the liquid. The yeasts that have been isolated from the mixture include Saccharomyces cerevisiae, Brettanomyces anomalus and Hansenula anomala.
The whole mixture is sometimes known as ‘Saccharomyces pyriformis‘ which is a synonym for Pichia membranaefaciens, a yeast that is available from many culture collections including the NCYC. However, although this yeast is often mentioned in connection with the ‘Bees Wine’, our records give no indication that it is a component of the mixture. Furthermore, any culture of Pichia membranaefaciens obtained from a culture collection would be a pure single yeast strain and would not give the same results as the mixture.
The gelatinous lumps formed in the ‘Bees Wine’ rise and fall as carbon dioxide is produced and released. Sugar, black treacle and ginger are fermented to produce a mildly alcoholic, rather sweet drink. Lemon/orange peel is sometimes added to improve the taste.
‘Bees Wine’ has several variations and is also known as ‘Ginger Beer Plant’, ‘Palestinian’ or ‘Californian’ Bees or ‘Balm of Gilead’.
The NCYC still keeps a culture of ‘Bees Wine’ in the laboratory although this is purely for scientific interest and none of the present staff have tried to make the ‘wine’ itself from the culture.
Due to health and safety considerations we are no longer able to supply the culture to the public as its exact composition is unknown to us.