Insect Control: Biological and Synthetic Agents - Index of

286 8: Mosquitocidal B. sphaericus: Toxins, Genetics, Mode of Action, Use, and Resistance Mechanisms
strains (Table 1) and are produced during sporulation.
The Mtx toxins are responsible for the activity
of most of the less active strains, and seem to be
synthesized only during the vegetative phase.
8.2.1. Crystal Toxins
The crystal toxin is composed of a 42-kDa protein
and a 51-kDa protein, hereafter designated BinA and
BinB, respectively (Baumann et al., 1987; Berry and
Hindley, 1987; Hindley and Berry, 1987; Arapinis
et al., 1988; Baumann et al., 1988; Hindley and
Berry, 1988; Berry et al., 1989). These two proteins
are synthesized in approximately equimolar
amounts, assembled in crystal structures and visible
at about stage III of sporulation (Yousten and Davidson,
1982; Kalfon et al., 1984; Baumann et al.,
1985). The genes encoding both proteins have been
cloned from several highly toxic strains. They encode
proteins with predicted molecular masses of 51.4
and 41.9 kDa (Baumann et al., 1987; Berry and
Hindley, 1987; Hindley and Berry, 1987; Arapinis
et al., 1988; Baumann et al., 1988; Hindley and
Berry, 1988; Berry et al., 1989). They appear to be
organized in an operon, with a 174- to 176-bp intergenic
region. A stem-loop structure (characteristic of
transcription terminators) has been identified downstream
from the binA gene (Figure 2, top; Hindley and
Berry, 1987; Baumann et al., 1988). No sequences
similar to B. subtilis sporulation promoters have
been found upstream from the binB gene. However,
both genes are expressed only during sporulation in
B. subtilis (Baumann and Baumann, 1989). Moreover,
lacZ fusion experiments have shown that
transcription begins immediately before the end of
exponential growth and continues into stationary
phase in B. sphaericus (Ahmed et al., 1995). This
probably allows sufficient amounts of the proteins
for crystal formation to accumulate by stage III.
In mutants of B. sphaericus 2362 strain in which
spo0A or spoIIAC sporulation genes have been
disrupted by insertional mutations, toxicity on
mosquito larvae is greatly reduced and no crystal
Figure 2 B. sphaericus toxin genes and schematic representation of corresponding polypeptides. Top, crystal (Bin) toxin: regions
shared by BinB and BinA are represented by identical colors. Inset: SDS-PAGE (PolyAcrylamide Gel Electrophoresis) of native
toxin and of toxin after activation by trypsin or by mosquito midgut juice. Bottom, Mtx toxins: the orange boxes indicate potential
signal sequences; the colors in Mtx1 represent the potential transmembrane sequence (red), the two regions shared with ADPribosyltransferase
toxins (green), and internal repeated sequences (yellow). Tryptic cleavage sites for each toxin are represented
by gray arrows. Potential ribosome binding sites (rbs) and terminator sequences (Ter) are also indicated.