Genetic, morphological, and virulence characterization of the entomopathogenic fungus Vericillium lecanii

Department of Agro-environmental Science, Obihiro University of Agriculture and Veterinary Medicine, Hokkaido, Obihiro 080-8555, Japan.
Journal of Invertebrate Pathology (Impact Factor: 2.11). 04/2003; 82(3):176-87. DOI: 10.1016/S0022-2011(03)00014-4
Source: PubMed


In order to clarify relationships among genetic diversity, virulence, and other characteristics of conidia, 46 isolates of Verticillium lecanii from various hosts and geographical locations were examined. The internal transcribed spacer (ITS) and intergenic spacer (IGS) regions of ribosomal DNA (rDNA), mitochondrial small subunit rDNA (mt-SrDNA) and beta-tubulin were analyzed by PCR-RFLP. PCR-single stranded conformational polymorphism (SSCP) was performed on regions of the mitochondrial large subunit rDNA, mt-SrDNA, beta-tubulin and histone 4. There were no relationships among the results of RFLP, SSCP, isolation source, and location. However, amplified product size of IGS did have relationships with conidia size and sporulation. Six isolates with 4.0-kb IGS products had large conidia dimensions, and yielded low numbers of conidia compared with other isolates. Three out of the six isolates were high virulence (over 90%) against green peach aphids. Furthermore, double-stranded RNA (dsRNA) was detected in 22 out of 35 V. lecanii isolates and related with the amplicon sizes of IGS, though not with virulence or isolation location. Isolates containing dsRNA were divided into six distinct types based on banding pattern. These data demonstrate the level of genetic diversity of V. lecanii, and suggest relations among the genetic properties and conidial morphology.

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Available from: Masanori Koike, May 09, 2014
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    • "Mitochondrial DNA (mtDNA), due to its properties to evolve faster than the nuclear DNA, to contain introns and mobile elements and to exhibit extensive polymorphisms, has been increasingly used to examine genetic diversity within fungal populations [24-26]. In other mitosporic entomopathogenic fungi, such as Metarhizium [27], Lecanicillium [28] and Nomurea [29], mtDNA data compared favourably to data based on ITS combined with a single nuclear gene, for applications in phylogeny, taxonomy and species or strain -identification. "
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    • "PCR-single strand conformation polymorphism, PCR-denaturing gradient gel electrophoresis, and PCR-temperature gradient gel electrophoresis SSCP (PCR-single strand conformation polymorphism ), DGGE (PCR-denaturing gradient gel electrophoresis ), and TGGE (PCR-temperature gradient gel electrophoresis) are used to detect genetic differences in PCR products obtained from SCL. SSCP relies on differences in secondary structure of single stranded DNA assessed by gel electrophoresis under non-denaturing conditions (Schwieger and Tebbe 1998), whereas DGGE and TGGE detect differences in DNA double-strand stability assessed by gel electrophoresis through a denaturant or temperature gradient (Muyzer and Smalla 1998). SSCP analysis have allowed discrimination of isolates of Lecanicillium lecanii (Ascomycota: Hypocreales) (Sugimoto et al. 2003), B. bassiana (Hegedus and Khachatourians 1996) or Nomuraea riley (Ascomycota: Hypocreales ) (Devi et al. 2007) by targeting loci coding for the mitochondrial small and large subunit rRNA, btubulin , or histon 4. DGGE has been used to discriminate B. bassiana isolates based on amplification products obtained from the ITS region (Pantou et al. 2003). TGGE has not been applied to entomopathogenic fungi, however it has a similar potential to SSCP or DGGE for identification of genotypes as demonstrated by studies on different yeast (Hernan- Gomez et al. 2000; Manzano et al. 2005) or bacterial species (Wagner-Dobler et al. 2000; Tominaga 2006). "
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    • "Lecanicillium lecanii ARSEF 6543, with homologies from 100% to 99% with T. confragosa isolates (IFO 8579 and UASWS 0012), appear close to some L. muscarium isolates, whereas the Mycotal isolate, identified as L. muscarium, appears in another different branch far from the rest (Fig. 1). Also Sugimoto et al. (2003) have found some similar ambiguities. All seems to indicate that the ITS region alone can fail to resolve some phylogenetic relationships, and is insufficient to place some isolates in their appropriate Lecanicillium branches in phylogenetic studies. "
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