卡森斯氏和短尾异小杆单菌线虫的制备方法

A method for culturing the entomopathogenic nematodes Steinernema kraussei and Heterorhabditis brevicaudis mononucleosis nematodes

本研究旨在建立一种利用昆虫病原线虫共生细菌培养单菌线虫的方法,可为检测线虫致病力和收集侵染期幼虫提供基础。首先分别分离了甘肃省抗低湿胁迫能力较强的卡森斯氏线虫(Steinernema kraussei) 0657L和非耐旱性短尾异小杆线虫(Heterorhabditis brevicaudis) 0641TY两个品系线虫的共生菌,然后在NBTB培养基上对分离到的共生细菌进行纯化和培养,随后将单菌落转移到LB培养基上。最后,从被侵染的大蜡螟(Galleria mellonella)体内收集种子怀卵成虫至培养有共生菌的LB上孵育,培养以收集侵染期线虫。研究结果表明,本研究所述方法简便快速地分离出了两个品系昆虫病原线虫的共生菌,并可通过培养共生菌收集侵染期线虫,筛选到一种直接从大蜡螟体内分离怀卵成虫至纯化的共生菌上孵育线虫,以快速准确获得整齐龄期的单菌线虫的简便方法。

The present study aimed to develop a method for culturing mononucleosis nematodes(MONO-nematodes) using the symbiotic bacteria(SB) of these nematodes, providing a basis for examining nematode pathogenicity and collecting infective juveniles(IJs). SB were isolated from two EPN strains collected from the Gansu Province: Steinernema kraussei,which is highly resistant to low-humidity stress, and Heterorhabditis brevicaudis, which is sensitive to low-humidity stress.The isolated bacteria were purified and cultured on NBTB. Subsequently, single colonies of SB were transferred to LB medium. Thereafter, the SB were incubated on LB medium with a gravid nematode female, which was collected from the body of an infected wax worm, and cultured to collect IJs. Therefore, the SB of S. kraussei 0657L and H. brevicaudis0641TY could be isolated using the method described here and IJs could be collected through culture with these SB. The proposed protocol can serve as a simple, rapid, and accurate method to collect IJs of MONO-nematodesdirectly isolating the gravid nematode females from infected wax worms and incubating them with purified SB.