Bt Research 2015, Vol.6, No.7, 1-3
1
A Letter
Open Access
Protocol for the fast isolation and identification of insecticidal
Bacillus thuringiensis
strains from soil
Palma L
1,2
1.Facultad de Ciencias Agrarias, Universidad Nacional del Litoral, Esperanza, Santa Fe, Argentina
2.Consejo Nacional de Investigaciones Científicas y Técnicas, Argentina
Corresponding author email
Bt Research, 2015, Vol.6, No.7
doi: 10.5376/bt.2015.06.0007
Received: 19 Aug., 2015
Accepted: 20 Sep., 2015
Published: 10 Oct., 2015
Copyright
©
2015
Palma et al., This is an open access article published under the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original work is properly cited.
Preferred citation for this article:
Palma L 2015, Protocol for the fast isolation and identification of insecticidal
Bacillus thuringiensis
strains from soil, Bt Research, Vol.6, No.7, 1
-
3 (doi:
Abstract
Bacillus thuringiensis
(Bt) is an ubiquitous, Gram-positive and sporulating bacterium that synthesizes insecticidal proteins
with specificity against a wide range of insects during sporulation (Cry and Cyt) and vegetative growth (Vip and Sip). These proteins
have portrayed Bt as an environment-friendly alternative to chemical insecticides. However, the intensive use of this resource has
promoted the evolution of insect resistance to some of the most used Bt strains to date. Continuous efforts are needed to obtain novel
strains exhibiting new specificities in order to overcome insect resistance and maintaining the insecticidal potential of this bacterium.
In this work, an experimental procedure for the fast isolation and identification of insecticidal Bt strains is described
.
Keywords
Bacillus thuringiensis
(Bt); Bt toxins; biological control
Introduction
Bacillus thuringiensis
(Bt) is a Gram-positive and
sporulated bacterium that synthesizes a number of
invertebrate active toxins with specific activity against
a wide range of insects (de Maagd et al., 2003;
Schnepf et al., 1998; van Frankenhuyzen, 2009). The
-endotoxins, including Crystal (Cry) and cytolytic
(Cyt) protein families are the best characterized group
of insecticidal proteins and are synthesized during the
stationary growth phase as crystalline parasporal
inclusions (Schnepf et al., 1998; van Frankenhuyzen,
2009). In addition, Bt also produces other insecticidal
proteins during the vegetative growth, the vegetative
insecticidal proteins (Vips) and the secreted insecticidal
protein (Sip), with toxicity against coleopteran (binary
Vip1/Vip2 and Sip toxins) and lepidopteran (Vip3)
pests (Donovan et al., 2006; Estruch et al., 1996;
Warren et al., 1998). Strains producing these insecticidal
proteins are commonly used in insecticidal formulations
and their encoding genes, in the construction of
transgenic plants (Sanchis, 2011), making the Bt-based
products, the most marketed microbial pesticides
worldwide (Roh et al., 2007; Schnepf et al., 1998).
The broad diversity of insecticidal genes encoded by
this bacterium strongly suggest that they resulted from
high selective evolutionary pressures (de Maagd et al.,
2003; Wu et al., 2007a; Wu et al., 2007b), that led
to the enlargement of their target ranges and made Bt
a broad source of insecticidal proteins (Palma et al.,
2014). On the other hand, insects targeted by Bt-based
insecticides, also suffered from selective pressures and
began to show symptoms of resistance to some of the
most frequently used insecticidal strains and toxins
(Ferré and Van Rie, 2002; Tabashnik et al., 2009).
These emerging insect resistance events are threatening
the efficiency of the most used Bt-based biopesticides
and demand the searching of novel insecticidal strains
exhibiting novel specificities and host ranges. This
protocol specifies the experimental procedure for the
fast isolation and identification of insecticidal Bt
strains from soil.
In this work, an experimental
procedure for the fast isolation and identification of
insecticidal Bt strains is described.
1 Sampling
In this protocol, soil samples were obtained with a
tubular soil sampler and consisted of ~20 g of
cultivated or non-cultivated soil after removing the
2-3 cm of the top layer, to prevent low abundance
levels of viable Bt spores due the effect of UV
radiation from sun (Iriarte et al., 1998). After
collection procedure, samples can be stored at 4 ºC in
