Detection and identification of baculovirus pesticides by multitemperature single-strand conformational polymorphism

• Published in: Journal of environmental science and health. Part B, Pesticides, food contaminants, and agricultural wastes Pesticides, food contaminants, and agricultural wastes, 2008-09, Vol.43 (7), p.539-545
• Main Authors: Szewczyk, Boguslaw, Barski, Piotr, Sihler, William, Rabalski, Lukasz, Skrzecz, Iwona, Hoyos-Carvajal, Liliana, Souza, Marlinda Lobo de
• Format: Article
• Language: English
• Subjects: Animals; Anticarsia gemmatalis; Baculoviridae; Baculoviridae - classification; Baculoviridae - genetics; Baculovirus; Biological and medical sciences; biopesticide; biopesticides; DNA primers; electrophoresis; Fundamental and applied biological sciences. Psychology; Genetic Variation; geographical variation; insect viruses; Insecta - virology; Lymantria dispar; Lymantria monacha; microbial detection; Nuclear polyhedrosis virus; nucleopolyhedrovirus; pathogen identification; Pest Control, Biological; Phytopathology. Animal pests. Plant and forest protection; Polymerase Chain Reaction; polymorphism; Polymorphism, Single-Stranded Conformational - genetics; Protozoa. Invertebrates; sequence amplification; single-stranded conformational polymorphism; Species Specificity; Spodoptera frugiperda; viral insecticides; Viral Structural Proteins - genetics
• ISSN: 0360-1234; 1532-4109
• DOI: 10.1080/03601230802234302

The method of single-strand conformational polymorphism (SSCP) was modified in our laboratories for the characterization of baculoviruses, insect viruses with great potential for use as bioinsecticides in biological protection programs. A series of primers were synthesized after the comparison of the polyhedrin gene sequences of over 20 baculoviruses. Polyhedrin is a highly conserved protein which is responsible for the persistence of the virus in the environment. Universal primers were designed which could be used in polymerase chain reactions (PCR) containing genomic DNA from an array of nucleopolyhedrosis viruses (NPVs) including these which are used as biopesticides against important pests of forests and crops, such as Anticarsia gemmatalis, Spodoptera frugiperda, Lymantria dispar, Lymantria monacha and many others. PCR products were denatured and subjected to single-strand DNA electrophoresis at variable temperatures (MSSCP) where, after silver staining, they gave ssDNA band patterns characteristic for each baculovirus species. This technique can be potentially applied to detect baculoviruses in insects collected in the field, as well as to plant tissues and the excrements or bodies of predators without need for sequencing the PCR products. Sometimes MSSCP can be used not only for species determination but also as an indication of genomic variability which can be related to infectivity.