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Appl Environ Microbiol. 2015 Aug;81(15):4894-903. doi: 10.1128/AEM.00671-15. Epub 2015 May 15.

Chromosome-Directed PCR-Based Detection and Quantification of Bacillus cereus Group Members with Focus on B. thuringiensis Serovar israelensis Active against Nematoceran Larvae.

Author information

1
Department of Microbiology, Swedish University of Agricultural Sciences, Uppsala, Sweden salome.schneider@slu.se.
2
Department of Environmental Science, Aarhus University, Roskilde, Denmark.
3
Department of Microbiology, Swedish University of Agricultural Sciences, Uppsala, Sweden.
4
Department of Medical Biochemistry and Microbiology (IMBIM), Zoonotic Science Center, Uppsala University, Uppsala, Sweden Department of Ecology and Genetics, Uppsala University, Uppsala, Sweden Swedish Biological Mosquito Control Project, Nedre Dalälvens Utvecklings AB, Gysinge, Sweden.

Abstract

Bacillus thuringiensis serovar israelensis is a wide-spread soil bacterium affiliated with the B. cereus group (Bcg) and is widely used in biocontrol products applied against mosquito and black fly larvae. For monitoring and quantification of applied B. thuringiensis serovar israelensis and its effect on indigenous B. thuringiensis serovar israelensis and Bcg assemblages, efficient and reliable tools are essential. The abundance and properties of B. thuringiensis serovar israelensis strains in the environment traditionally have been investigated with cultivation-dependent techniques, which are hampered by low sensitivity and the morphological similarity between B. cereus and B. thuringiensis. Currently available PCR-based detection and quantification tools target markers located on plasmids. In this study, a new cultivation-independent PCR-based method for efficient and specific quantification of B. thuringiensis serovar israelensis and Bcg is presented, utilizing two sets of PCR primers targeting the bacterial chromosome. Sequence database searches and empirical tests performed on target and nontarget species, as well as on bulk soil DNA samples, demonstrated that this diagnostic tool is specific for B. thuringiensis serovar israelensis and Bcg. The method will be useful for comparisons of Bcg and B. thuringiensis serovar israelensis abundances in the same samples. Moreover, the effect of B. thuringiensis serovar israelensis-based insecticide application on the total Bcg assemblages, including indigenous populations, can be investigated. This type of information is valuable in risk assessment and policy making for use of B. thuringiensis serovar israelensis in the environment.

PMID:
25979887
PMCID:
PMC4495189
DOI:
10.1128/AEM.00671-15
[Indexed for MEDLINE]
Free PMC Article

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