Characterization And Pathogenicity Research Of Entomopathogenic Nematode 09FLY1 From Pristionchus Genus

Entomopathogenic nematodes (EPN) are promising biological control agents since the early of 20th century. Entomopathogenic nematodes are obligate and lethal to a wide range of insect pests, active search parasitifer, safety for human and animals, without contamination, don't produce resistance and can be applied with conventional pesticide equipment or other factor of biocontrol. So, entomopathogenic nematodes have been widely used to control pest insects in agriculture, and attached importance to many researchers and commercial departments. Up to today, there are 78 valid EPN species:14 species have been reported in the genus Heterorhabditis of family Heterorhabditidae,62 valid species in the Steinernema and 1 species in the Neosteinernema of family Steinernematidae, and 1 species in the genus Heterorhabditidoides of Rhabditidae. All of the described species of EPN are belong to Rhabditida.The new EPN strain 09FLY1 was obtained from a soil sample collected from Yixing Jiangsu province using the insect baiting method. Body length 260-900μm, cuticle annulated and longitudinal ridge, hermaphrodite, degradation of masculine characteristics,eurystomatous, esophagus present and long, two bulbs median bulb and extrem bulb present, Six teeth in the metrhabdion, two amphids on the cephalic six papillaes and flabs, trail present and slender, vulva without cuticular flaps to protected. Infective juveniles:body length=244.30±11.33 (228.82-285.19), pharyngeal length= 37.846±1.154 (33.27-53.82), two bulb distance=18.463±1.021 (17.34-29.11), tail length=101.307±14.706 (98.84-132.19), ES=43.431±3.123 (40.85-61.30). Molecular phylogenetic trees based on 18S rDNA revealed that 09FLY1 was belong to the genus Pristionchus and has a closer relationship with Pristionchus pacificus. The percentages and length of the nucleotides A, T, C and G in the 18S rDNA and ITS regions of 09FLY1 are significantly difference from that of Pristionchus pacificus and other species of the genus Pristionchus. Morphological characteristics of the nematode combined with molecular data showed that the nematode 09FLY1 is a new species of the genus Pristionchus. The mortality of Galleria mellonella larvae was 94% when infected by the infective juveniles of 09FLY1 in 4 days. The value of LD50 to Galleria mellonella larvae was 14 nematodes after 6 days.A novel entomopathogenic symbiotic bacteria strain 09FLYB1 was isolated from the intestine of 09FLY1.Comparative 16S rDNA sequences analysis indicated that the bacteria is a members of the genus Serratia.The pathogenicity of the strain 09FLYB1 to Galleria mellonella larvae and C. zealandica larvae was tested. The disease symptoms of C. zealandica larval include cessation of feeding, amber coloration, and eventual death. The bacteria strain had significant pathogenic to Galleria mellonella larvae, and the LD50 was 86 bacteria cells.