Resistance And Genetic Differentiation Of Rhyzopertha Dominica To PH3at Different Spatial Scales In China

Rhyzopertha dominica (Fabricius) is one of the major coleopterous pests of stored products in China and around the world. Both larvae and adults are internal feeders, causing serious damages to grains, such as rice, maize, wheat and other stored commodities. It causes serious losses to stored grains in tropical and subtropical region. Currently, the main method for controlling stored-product insects in many countries, including China, is fumigation with phosphine. However, repeated ineffective fumigation in poorly sealed structures resulting in under-dosing could lead to the development of strong resistance in target pest. This research investigated the phosphine resistacne and the instantaneous growth rate (ri) among different geographical populations of R. dominica in China's grain region. AFLP and mtCOI gene also used to analyze the genetic differentiation among R. dominica population. The activities of catalase (CAT), superoxide dismutase (SOD) and peroxidase (POD) were also observed (CAT, SOD, POD). And the detail results showed as followed:1. Phosphine resistance in R. dominica in China from different geographical populations in ChinaThe present investigation was carried out to determine the level of phosphine resistance in16R. dominica (Fabricius) populations that were collected from ten provinces and one municipality in China following the Food and Agriculture Organization's (FAO) standard method. Results showed that the50%lethal concentration (LC5o) of phosphine to these R. dominica populations ranged from0.017mg/L to4.272mg/L. Of the16populations,5were of low resistance,6were moderately resistant, and5were high resistant. The present results have revealed the seriousness of the problems of phosphine resistance in China. R. dominica populations with high ri showed low resistance factor (RF for LC50), while low ri presented high RF value. The populations were ranked in order of RF values, the populations from processing facilities cluster near the top, then the national storage populations, then the populations from local storage. The average RF for processing facilities, national storages and local storage are5.465,61.490and277.643, respectively. This result indicated that resistance is connected with the type of grain storage.2. Genetic diversity among different geographical populations of R. dominica in China as revealed by AFLP In this study, the genetic diversity and structure of16populations (110individuals) of R. dominica were examined using Amplified Fragment Length Polymorphism (AFLP) analysis. Four primer combinations were selected to identify a total of128polymorphic loci ranging from50to450bp. The percentage of polymorphic loci (Pp) was43.95%at the population level and84.87%at the species level, respectively. Nei's gene diversity (h) and Shannon's Information index (I) had similar trend with the percentage of polymorphism. Thus, the estimated genetic diversity using these three indicators revealed a high level of genetic variation within each R. dominica population. At the species level, Analysis of Molecular Variance (AMOVA) revealed a significantly greater genetic variation within populations (82.38%) than among populations (17.62%)(P<0.0005). This result showed that the genetic diversity mainly occurred within the population, not among populations. The ΦST value of the specied was0.176and the gene flow (Nm) value was1.170at the species level. This indicated that, a certain level of gene flow was among the populations and prevented the genetic differentiation among populations caused by genetic drift.An Unweighted Pair Group Method with Arithmetic Mean (UPGMA) dendrogram based on Nei's unbiased genetic distance revealed four distinct groups that did not correspond to the geographical regions. The Mantel test of isolation by distance revealed that there was no significant correlation between Nei's genetic distance and geographical distance (Lg) among all populations (r=0.1054, P=0.8034>0.05). This result suggests that isolation by distance does not describe the pattern of gene flow among R. dominica populations. The ecology of grain storage area and the depot/storage facility type were not the main factors determining the genetic diversity of R. dominica.3. The analysis of mtCOI gene and genetic variation of R. dominica populationThe mtCOI gene partial sequences (691bp) of100individual from15R. dominica populations were amplified and sequenced. Nucleotides compositi on, transiti on and transversi on, genetic distance of this segment had been analyzed. The molecular volution characteristic of COI gene in different individuals was discussed. The results indicate that average A+T content of COI gene in R. dominica is63.2%, which shows strong A+T bias. The A+T content was85.2%,53.3%and57.0%respectively in the three points. A+T content of this bias showed stronger in the first than that of the second and third points. A, T, C, G content of33.2%,32%,18.7%,16.2%. Twenty-six nucleoide site were substituted and twenty-five haplotypes were identified in all sequences with seven shared. The Nm value was1.35showed that there was certain gene flow among R. dominica population. AMOVA analysis of genetic structure indicated that the gentic variation of R. dominica mainly occurred among individuals.4. Effect of phosphine on three protective enzymes in R. dominica (Fabricius) adult in different geographical populationsThe activities of catalase (CAT), superoxide dismutase (SOD) and peroxidase (POD) were observed in the R. dominica adult in seven different geographical population. The activities of CAT, POD and SOD showed difference among populations in R. dominica adults before fumigation. The results showed that the activities of CAT and POD in seven different population of adults decreased after fumigation treatment, While the activities of SOD enhanced. The activity of CAT in seven populations had expressed significant simple correlation with the50%lethal concentration (LC50) of R. dominica populations (P=0.031<0.05); Meanwhile, there were no significant correlation between the SOD, POD and the LC50three enzyme activities before fumigation (P=0.545;0.053>0.05), while the CAT activity showed simple correlation with the POD activity (P=0.034<0.05).