Genomic DNA extraction methods and phylogenetic analysis of Beauveria bassiana from Central Java, Indonesia, and its toxicity against the fall armyworm, Spodoptera frugiperda J.E. Smith (Lepidoptera: Noctuidae)

Abstract

Abstract Background Control techniques using biological control agents such as Beauveria bassiana (Balsamo) Vuillemin have the advantage of not showing any negative impacts on environmental health and safety issues. This study used isolates from B. bassiana collection from Laboratory of Pest Monitoring belonging to Indonesian Ministry of Agriculture (LPHP) in Central Java which showed potential in controlling target pest. The problems that still occur are the lack of facilities and infrastructure and the lack of quality testing of collection isolates in LPHP, so that the isolate identification process is still carried out in simple method, and bioassay testing on the fall armyworm (FAW), Spodoptera frugiperda J.E. Smith (Lepidoptera: Noctuidae) as a target pest, is not commonly conducted. The results of bioassay testing can be used to determine the potential of a biological agent to control target pest. Result The two DNA extraction methods showed different results regarding DNA concentration and purity values, but both methods were good and could be used to amplify DNA using PCR. The DNA band was amplified at 500–600 bp using primers ITS 1 and ITS 4. The results of molecular analysis showed that the four isolates of B. bassiana from Central Java were found in the same clade as B. bassiana from South Sumatra, Dhaka, and Oromia, where these isolates showed similar similarities descended from a common ancestor. Genetically, B. bassiana isolates from Central Java show more genetic similarities to B. bassiana isolates from South Sumatra, Indonesia. Quality testing was carried out by calculating the density and germination ability values for LPHP isolates from Sukoharjo (Sukoharjo isolate), Temanggung (Purworejo isolate), and Banyumas (Banyumas and Cilacap isolate), which showed varying results. The bioassay test used three isolates, namely B. bassiana from Sukoharjo, Banyumas, and Cilacap, which were selected based on density values, germination ability, and molecular analysis. The ability to cause death of the three isolates against S. frugiperda showed different results where the isolate from Sukoharjo, Banyumas, and Cilacap caused mortality of 60, 40, and 60%, and the LC50 value of each isolate was 3.3 × 106, 1.3 × 107, and 3.5 × 107 conidia ml−1, respectively. Conclusion Morphological identification by macroscopic, microscopic, and molecular analysis showed that the isolate from the LPHP collection in Central Java, Indonesia, was B. bassiana. Genetically, the four isolates showed similar characteristics to isolates from South Sumatra, Indonesia. B. bassiana isolates from collections from Central Java showed potentials as a biological control agent against S. frugiperda.

Keywords

• Phylogenetic analysis
• Beauveria bassiana
• Kit extraction
• CTAB
• Kimura two-parameter
• BIC