Identification Of The Novel Entomopathogenic Nematodes Species RGO81015 Of The Genus Heterorhabditidoides And Its Symbiotic Bacteria Strains DR186 And R187

Entomopathogenic nematodes are soil-inhabiting, lethal insect parasites that have been developed commercially as a biocontrol agent and successful. EPNs include Steinernem-atidae, Neosteinernema, Heterorhabditis and Heterorhabditidoides. Steinernematidae and Neosteinernema belong to Steinernematidae. Heterorhabditis belongs to Heterorhabditidae. Heterorhabditidoides belong to Rhabditidae.all of them belong to Rhabditida, Secernentea, Nematoda,animality. Steinernematids Heterorhabditids and Heterorhabditidoides carry specific symbiotic bacteria of the genus Xenorhabdus, Photorhabdus and Serratia nema-todiphila in their intestine characteristic.The new species was obtained from a soil sample collected from a kailyard of Rugao City, Province of JiangSu by the method of insect baiting, designate RG081015. For males: Body length= 1299.65(1143.00-1474.49)μm. D=66.6(20-51.09)μm; ES=19.53(15.05-25.74)μm; Spicule length=49.23(35.24-60.93)μm; Bursa like a butterfly, well developed with nine pairs of genital papillae, the papillae formula of bursa is 1,2,3,3.The number and arrangement don't change, a pair of Spicules separate Symmetri- cally. Female:Body length=1401.36 (1117-1588)μm; D%=941.89(787-1105)μm; Surface has four distinct lateral line; Bare vulva. Infective juveniles, body length= 622.01(390-863)μm; EP=160.89(135-200)μm; tail length with sheath =90.69(70-107)μm; D%=859.13 (815.5-901.1)μm; E%=25.83(23.49-28.5). The morphological characteristics of this nematode is similar to that of H. chongmingensis, but there are lots of obvious different. For instance, compared with H. chongmingensis DZ0503CMFT, the male of H. chongmingensis DZ0503CMFT's labial congstrict, but that phemomenon are not so clearly for RG081015, a ligulate in RG081015's pharynx, the intine of oral cavity keratinize obviously, the emago of RG081015's body and tail length is longer(about 184μm),the oral cavity is longer and narrower, the excretory pore is closer with foreside of the body, the spicule of RG081015 is more developed. The 18S rDNA gene sequence have 2 sites difference,homology 99%,ITS sequence have 12 sites difference, homology 98%. The Molecular phylogenetic trees based on 18S rDNA and the internal transcribed spacer sequences data revealed that RG081015 and H. chongmingensis DZ0503CMFT are in the same group. They are in the same genera of Rhabditidae. The cross breedings are fail in the two nematodes,what indicate that the reproductive isolation exist between them.That all testify the two species nematodes are different from eath other.Two strain bacteria are obtained from 66 single colony isolated from the intestine of nematoda RG081015, which are red-pigmented, Gram-negative, designate DR186 and R187. Molecular phylogenetic trees based on 16S rDNA and the rpoB gene sequences data revealed that both DR186 and R187 belong to Serratia,the symbiotic bacteria of RG081015 and H. chongmingensis DZ0503CMFT pool in one group, they classify in the same genus. DR186 and R187 are different from DZ0503SBS1T in Double-arginine hydrolase activity, Amylase activity, Nitrate reductase,Tween 80 hydrolysis activity. DR186 and R187 can't use D-melibiose, starch,but DZ0503SBS1T can. DR186 and R187 are insensiti- ve to Ampicillin, erythromycin, penicillin G, rifampin,but DZ0503SBS1T is sensitive.