Isolation Of The Yeast-like Symbiont Pichia Anomala(Hemiascomycetes: Saccharomycetaceae) From Laodelphax Striatellus(Hemiptera: Delphacidae) And Its Sensitivity To Fungicides

Small brown planthopper Laodelphax striatellus is one of major agricultural pests and widespread in C hina. It is not only sucking rice, barley, wheat and other gramineous crops,but also spreading kinds of crop virus. The utilization of chemical pesticides was used as main method for control of L. striatellus. While, with the extensive application of chemical pesticides, the L. striatellus’ s has increased its resistance to pesticides, so control effect of pesticides on L. striatellus was weakened and environmental pollution was worsen. Therefore it is crucial to explore a new method for controlling this pest. Yeast-like symbiontes(YLS) are endosymbionts that are closely linked with the growth, development, virulence variation and pesticide resistance of their host. ―Controlling pests through inhibiting YLS‖(CPY) provide a new idea for L. striatellus’ s control. The core idea of CPY was that inhibiting the quantity of symbiontes inside of L. striatellus was to weaken the vitality and resistance of L. striatellus, and to improve the control effect of the pesticide. In this study, the YLS was isolated from L. striatellus via in vitro culture and the susceptibility of isolated YLS to different concentrations of fungicides was examined. Selection of appropriate fungicides and its effects on CPY were also determined. The results were as follows:1. Isolation and identification of YLS in L. striatellus: a strain of YLS was successfully isolated and identified as Pichia anomala based on corresponding carbon assimilation experiment and 18 S r DNA phylogenic analysis.2. Confirmation of the existence and quantity of P. anomala: the denaturing gradient gel electrophoresis(DGGE) was employed to analysis the diversity of YLS and confirm the existence of isolated strain P. anomala in L. striatellus. The fluorescent in situ hybridization(FISH) result further confirmed that a certain number of P. anomala exist in L. striatellus’ s abdomen. Subsequently, the variation and copy number of P. anomala in different L. striatellus instars was measured by using Real-time florescent quantitative PC R(q PCR). Under the different nymphal instars of L. striatellus, the number of P. anomala gradually increased with the increase of nymphal instar of L. striatellus until the 5t h instar, and the number of P. anomala decreased significantly in the 5t h instar and then increased rapidly in the newly-emerged female adult.3. Influence of different pesticides on L. striatellus and its YLS: The susceptibility of P. anomala to different concentrations of fungicides was tested. The results indicated that 70% propineb had obvious inhibitory effects on the growth of P. anomala, the 50% tebuconazole+25% trifloxystrobin next, while the fluopicolide+ propamocarb hydrochloride and 40% pyrimethanil had relatively worse effect; When two fungicides(70% propineb and 50% tebuconazole +25% trifloxystrobin) with obvious inhibitory effect were applied on the wheat seedlings at the concentration of 800 mg/L, the mortality of L. striatellus adults feeding the treated wheat seedlings reached 46.7% and 63.3%, respectively, which was apparently higher than that of the control group, and the body weight of L. striatellus adults in treatment group was also less than that of the control group. Real-time florescent quantitative PCR results indicated that the numbers of YLS of three strains(Hypomyces chrysospermus, Pichia guilliermondii, and Pichia anomala) inside L. striatellus dropped sharply after different fungicides were used. The mortality of L. striatellus was negatively correlated with the quantity of YLS in trend.4. Influence of blended pesticide to L. striatellus’ s control: the 70% imidacloprid with 70% propineb and 50% tebuconazole +25% trifloxystrobin was mixed, respectively. The virulence of 70% imidacloprid+ 1 600 mg/L(50% tebuconazole +25% trifloxystrobin) on L. striatellus was enhanced obviously comparing to that of 70%imidacloprid, while 70% imidacloprid+ 1 600 mg/L 70% propineb has a minor improvement of virulence on L. striatellus. When 10 mg/L 70% imidacloprid mixed with 1 600 mg/L(50% tebuconazole +25% trifloxystrobin) as 1:1 was used, the mortality of L. striatellus were significantly higher than that of 70% imidacloprid in corresponding concentration.