Safety Evaluation Of Xenorhabdus Nematophila On Non-target Organisms

Xenorhabdus nematophila HB310 strain,an entomopathogenic bacterium symbiotically associated with entomopathogenic nematode Steinernenza carpocapsae,appeared high oral toxic activity against several species of insect.In order to explore the feasibility of the X.nematophila HB310 strain as bioinsecticidal agent or transgenic crops,the objective of study determined the biological safety of X.nematophila HB310 culture broth and the insecticidal protein on non-target organisms,including Trichogramma,Microplitis mediator,Harmonia axyridis and Bombyx mori. Effects of X.nematophila HB310 to the biological diversity in cabbage field were systematically evaluated at the same time.The study will provide the basis for the utilization of X.nematophila HB310 in the future.The main results are as follows:1.No effect was observed on the emergence rate of Trichogramma chilonis and Trichogramma brassicae treated with HB310 culture broth and insecticidal protein crude extracts.The longevity of adult of the two kinds of Trichogramma feeding on the honey with HB310 culture broth,cells and supernatant were shorter than control,and the longevity of adult got shoter with increase of the concentration.If the concentration of toxinâ…¡did exceed lμg/mL limit,the toxinâ…¡would not influence Trichogramma adult.2.No obvious effect was observed on the emergence of Microplitis mediator after treating cocoons with HB310 culture broth and insecticidal protein crude extracts.The longevity of the M. mediator adult feeding on the honey with HB310 culture broth and insecticidal protein crude extracts was shortened.But when the concentration of HB310 insecticidal protein extracts reduced to 100μg/mL,it did not influence the longevity of M.mediator adult.Feeding on the honey with 20μg/mL toxinâ…¡,the longevity of Microplitis mediator adults were decreased significantly.No detrimental effect was observed on the growth duration,cocoon weight,cocconing rate and emergence rate of M.mediator after host larvae feeding on X.nematophila HB310 culture broth and insecticidal protein crude extracts.Compared with the control,the larval duration of M.mediator was delayed significantly when the host larvae fed on diet containing 227.04μg/mL HB310 insecticidal protein extract.3.The predatory function of Harmonia axyridis was not affected by HB310 culture broth and insecticidal protein crude extracts when they fed on Myzus persicae dip in toxin or Plutella xylostella fed on toxin-diet.The of exposure of HB310 insecticidal protein crude extract(10.00μg/mL)in short time(just at 72 h)or the low concentration showed less different compared with control H.axyridis in growth,pupation rate,pupa weight,emergence rata and survival rate.On the development and other fitness parameters,there were no significantly differences between H.axyridis predating toxin-treated P.xylostella larvae and control-treated P.xylostella larvae.But the pupation rate and survival rate(at 15 d)of H.axyridis predating P.xylostella larvae which previously fed on HB310 culture broth (3.73×10³ CFU/mL)were decreased significantly.4.X.nematophila HB310 showed less toxicity to the silkworm.The acute oral toxicity of HB310 culture broth was too low to detected out LC₅₀to silkworm larvae even at 120 h after toxin-treatment (LC₅₀＞2.24×10⁶ CFU/mL),but high concentration of HB310 culture broth and insecticidal protein crude extracts all showed obvious antifeeding activities and growth inhibition to silkworm larvae, even affected to cocoon weight,cocoon shell weight,pupa weight and pupation rate of silkworm.The antifeedant rates of culture broth(2.24×10⁶ CFU/mL)and insecticidal protein crude extract (567.60μg/mL)to 5^thinstar larvae was 24.90%and 23.05%respectively.The larvae with toxin-feeding treatment in short time(just at 72h)or the low concentration of HB310 culture broth (1.12×10⁴ CFU/mL)and insecticidal protein crude extracts(113.52μg/mL)showed less difference compared with control larvae in growth and development of 3^th-5^thinstar larvae,pupation rate, cocoon weight,pupa weight,cocoon shell weight,emergence rata and fecundity of adult.5.Systematic surveys were implemented on effect of X.nematophila HB310 on population dynamics of the main pests,enemies and non-target organisms in the cabbage field.The insecticidal activity of X.nematophila HB310 to P.xylostella(49.43±22.05%on Jun.6)had no obvious differences compared with that of Bacillus thuringiensis HD-1(59.24±20.05%).And the persistence of HB310 culture broth to Pieris rapae was shorter than that ofBt HD-1,(HB310 effect 52.10±2.33%, while HD-1 72.73±1.60%).HB310 did not showed obvious influence to diversity of arthropod community.