Insect Control: Biological and Synthetic Agents - Index of

A8 Addendum: Bacillus sphaericus Taxonomy and
Genetics
C Berry, Cardiff University Park Place, Wales, UK
M H N L Silva Filha, Centro de Pesquisas Aggeu
Magalha˜es-Fundac¸ ão Oswaldo Cruz, Recife-PE,
Brazil
ß 2010 Elsevier B.V. All Rights Reserved
A8.1. Mtx1 Toxin Structure 308
A8.2. New Toxins 308
A8.3. Characterization of Other Bin Toxin Receptors in Culicide Larvae 309
A8.4. Characterization of Resistance Alleles and Resistance Management 309
Although still widely referred to as Bacillus sphaericus,
a taxonomic revision has redesignated this
bacterium as Lysinibacillus sphaericus (Ahmed
et al., 2007). Our understanding of this bacterium
has been enhanced by genome sequencing of the
mosquitocidal serotype H5a,5b strain C3-41 (Hu
et al., 2008). The genome revealed a lack of important
enzymes and transport systems that explain the
inability of this species to metabolize sugars. A new
gene encoding an Mtx2/Mtx3-like protein (Mtx4)
was also identified along with a further pseudogene
in this family that is located upstream of the mtx3
gene. This implies that recombination may be a
feature of this protein family and all the genes in
this family appear to be associated with mobile
genetic elements (Hu et al., 2008). The mtx2 gene
in strain C3-41 lies close to the mtx1 gene and the
mtx4 gene lies close upstream of the bin genes, but
other toxin genes are distributed around the genome.
It is of interest to note that in this strain,
mtx1 contains a stop codon and appears, therefore,
to be a pseudogene. The bin genes encoding BinA
and BinB toxins are present along with the mtx4
gene on an 35 kb region that is duplicated in the
genome and on a large plasmid pBsph. The existence
of such duplication had been suggested earlier for
strain 2297 (Poncet et al., 1997).
A8.1. Mtx1 Toxin Structure
The structure and activity of the Mtx1 toxin has
been studied in some detail (reviewed in Carpusca
et al., 2006). The 100 kDa protein is processed in
the insect gut to a 27 kDa fragment with ADPribosylating
activity and a 70 kDa fragment thought
to mediate entry of the 27 kDa portion into target
cells (Thanabalu et al., 1992, 1993; Schirmer et al.,
2002). ADP-ribosylation of target proteins in mosquito
cells and in other eukaryotic and bacterial
cells (Schirmer et al., 2002) has been reported
and appears to occur by modification of arginine
residues (Schirmer et al., 2002). Enzymatic activity
of the activated 27 kDa protein can be inhibited by
the binding of the 70 kDa protein and a 20 amino
acid region responsible for this effect by competition
with NAD + (Carpusca et al., 2004). The C-terminal
70 kDa fragment contains four QxW3 repeats,
characteristic of ricin toxin lectin-like sequence
motifs (Hazes and Read, 1995). A further, noncompetitive
inhibition of ADP-ribosyl transferase activity
by these lectin domains has also been reported
(Carpusca et al., 2004). Crystal structures for the
N-terminal region (Reinert et al., 2006) and the
holotoxin (Treiber et al., 2008) illustrate many of
the features of the toxin including auto-inhibition
and activation sites.
A8.2. New Toxins
In addition to identification of the mtx4 gene mentioned
earlier, other toxins have been identified recently
in B. sphaericus strains. Sphaericolysin is a
53 kDa-secreted protein that, on injection, shows
insecticidal activity against the German cockroach,
Blattela germanica and to a lesser degree, against
Spodoptera litura (Nishiwaki et al., 2007). Originally
isolated from strain A3-2, it is also encoded by
the genome of strain C3-41 (Hu et al., 2008). Strain
A3-2 was isolated from the ant lion Myrmeleon
bore, which may use the toxins to kill its prey
(Nishiwaki et al., 2007). Sphaericolysin appears to
be a member of the cholesterol-dependent cytolysins
and causes neuronal damage in intoxicated insects.