A formal identification will place the fun-
gal isolate within a taxonomic rank, initially at
species level. Many fungal species are widely
distributed in the environment, and therefore
there is also a need to be able to identify the
individual isolate that is under consideration.
This can be undertaken through techniques,
such as DNA fingerprinting (Caetano-Anolles
et al., 1991; Schlick et al., 1994; Bridge et al.,
1997), which allow the identification of either
single strains or groups of very closely related
strains. This level of identification is particu-
larly important as it provides a mechanism for
tracking the progress and fate of the agent in
the environment, a validation check for the
purity and accuracy of the formulations and a
standard reference that may be used to regis-
ter or protect individual isolates (Schlick et al.,
1994; Edel, 1998; McClintock, 1999; Neal and
Newton, 1999).
It has long been accepted that serial sub-
culturing of fungal isolates on agar can result
in the loss of certain characteristics of the orig-
inal isolate (Thomas and Smith, 1994). This
can be avoided by careful management of
fungal material from the point of original iso-
lation. Once a clean, uncontaminated culture
has been obtained, subcultures from this
should be made and stored, using as many
long-term storage methods as possible. Two
methods that are commonly used by large
culture collections are lyophilization (freeze-
drying), cryopreservation (using liquid nitro-
gen) and storage at 80°C. All of these
techniques require the purchase of relatively
expensive equipment but can be contracted
out to culture-collection laboratories with
experience in their use. Lyophilization or
freeze-drying can be very useful as a large
number of freeze-dried ampoules can be pre-
pared and stored long-term (10 years) at
room temperature (Smith and Kolkowski,
1996). Liquid nitrogen or cryopreservation is
also an effective method of long-term storage,
but maintenance of cultures preserved using
this technique requires a regular supply of liq-
uid nitrogen and highly specialized preserva-
tion tanks. Cultures preserved using this
method are therefore maintained at the site of
preservation and must be requested from
storage each time that a source of inoculum is
required. Storage at 80°C requires a special-
ized freezer, but may be the most economical
option for small companies interested in
maintaining their own culture collection.
Techniques requiring less sophisticated equip-
ment can also be effective and can be used as
a backup to the above techniques. These
include silica-gel storage, storage under min-
eral oil, soil storage and water storage (Smith
and Kolkowski, 1996). Some of these tech-
niques are not suitable for all types of fungi.
The most appropriate method/methods
should be selected for each organism to be
preserved. In preparation for mass produc-
tion, a large stock of fungal material should
be prepared and stored using a suitable long-
term storage method. Fresh starter material
for each production run should then be pre-
pared, using material direct from the stock.
Recent studies on long-term preservation
techniques have uncovered some worrying
effects on both phenotypic characteristics and
genetic stability of some of the fungal isolates
studied (Ryan, 1999). Detailed studies on a
number of mitosporic fungi have shown that
both lyophilization and cryopreservation can
cause detectable molecular polymorphisms.
This was particularly marked if generalized
protocols (as opposed to one that has been
optimized for the particular fungus in ques-
tion) were used for preservation (Ryan, 1999).
This highlights the need for careful consider-
ation before placing valuable isolates in long-
term storage. In response to this, Ryan et al.
(2000) have developed a key for use in the
selection of fungal preservation techniques.
One safeguard against the potential loss
of valuable characteristics of some mycopes-
ticides (particularly mycoinsecticides) is the
regular passaging of the fungal isolate
through the host or closely related species.
However, this process should be carefully
controlled to ensure that the population used
for passaging is not harbouring latent infec-
tions, which may result in cross-contamina-
tion with or isolation of the wrong pathogen.Production process and record keepingProduction processes, downstream process-
ing (harvest and postharvest) and extraction
methods vary widely from product to prod-Quality Control of Fungal and Viral Biocontrol Agents 249