turing agents, spreaders, stickers and stabi-
lizing agents (Burges, 1998). Additives such
as wetting agents and gustatory stimulants –
for example, molasses or crude sugar – may
be introduced to tank mixes prior to spray-
ing to improve efficacy and persistence
(Burges and Jones, 1998).
The process of in vivoBV production is
conceptually very simple and can require
minimal equipment. In practice, maintaining
a sustained production of virus is demanding
and it is only with well-developed produc-
tion procedures and a rigorous standard of
process quality control that effective produc-
tion can be both attained and maintained.
Culture maintenanceQuality control in BV production starts with
the raw materials, which, for in vivoproduc-
tion, include not only the pathogen but also
the live host insects. The virus used for inoc-
ula should be a properly identified strain
that has been characterized by DNA restric-
tion analysis. The inoculum should be puri-
fied using gradient centrifugation (Hunter
Fujita et al., 1998) to ensure that it is virtually
free of contaminating insect pathogens,
which could infect the hosts used for pro-
duction and interfere with viral replication,
reducing BV productivity. One practice that
should always be avoided is to use recycled
BV from production as inoculum for new
batches without characterization or purifica-
tion. This is a common practice in some low-
technology NPV-production systems and can
be a major cause of problems.
The insect cultures used for BV production
should be free of parasites and monitored for
the presence of latent virus infections. For this
reason, the preferred hosts for production are
disease-free laboratory insects reared under
controlled conditions and on a standardized
artificial diet. The use of laboratory-reared
insects ensures that larvae can be infected at
the right developmental stage for optimum
productivity. Several studies have shown that
optimizing the larval age/weight at infection
is valuable in maximizing BV productivity in
individual larvae (Cherry et al., 1997;
Grzywacz et al., 1998).
Ideally, detailed records of egg laying,
hatching rates, pupal size and larval/pupal
mortality rates should be maintained, as well
as rates of pupal deformation (Moore et al.,
1985). These data should be plotted on
graphs and scrutinized at regular intervals,
as deviations from norms are often the best
early indicator of insect health problems. It
can also be very useful to carry out regular
weekly microscopic examinations on any
‘sickly’ insects and a few randomly selected
ones to check for possible infection, using a
standard guide to entomopathogens, such as
that of Poinar and Thomas (1984).
Some insects appear to adapt very readily
to culture and can survive for many (50)
generations without the need to introduce
new genetic stock; Spodoptera littoralisis a
good example of this (McKinley et al., 1984).
Such cultures are relatively easy to maintain
and are a valuable asset, as the introduction
of new insects is the commonest route by
which unwanted infections enter a culture.
However, laboratory cultures of other
species, e.g. H. armigera, have been observed
to decline in vitality after a number of gener-
ations and require regular introductions of
new insects. Where insects are to be intro-
duced into established cultures, rigorous
checking of the new insects is important. The
new stock should first be held in a separate
quarantine unit. Precautions should include
egg washing, rearing in pairs or small
groups and keeping eggs separate to mini-
mize the spread of infection. Combined with
close examinations of larvae for signs of ill
health and microscopic examination of
females for infection (after they have fin-
ished egg laying), these precautions can be
used to eliminate contaminated breeding
lines (Chapter 10).
The design of the insect production facil-
ity needs to be carefully planned to mini-
mize the danger of the colony becoming
infected by either the production BV isolate
or any wild pathogens. Examples of
designs for appropriate facilities are avail-
able in the literature (Bell et al., 1981;
Hunter-Fujita et al., 1998) and published
protocols and methodologies for mass-rear-
ing host insects are available (e.g. Singh
and Moore, 1985).Quality Control of Fungal and Viral Biocontrol Agents 255