RESUMO
Rhyzopertha dominica Prip (RdPrip) cDNA was cloned (GenBank accession no. OK318454), and the encoded 276-amino-acid protein indicated the typical aquaporin structure, including six transmembrane regions and two NPA motifs. The developmental and tissue profiles of RdPrip transcription were determined. RdPrip was highly transcribed in female adults, followed by larvae, pupae, and male adults. The transcriptional expression levels of RdPrip were significantly high in the ovary and hindgut (including cryptonephridial systems) compared with the foregut, testis, midgut, and Malpighian tubules. Knockdown of RdPrip in female adults did not decrease fecundity, but significantly decreased the hatching rate of eggs laid by the females. The results suggest that RdPrip functions in embryonic development, not in egg formation. In addition, the transcriptional expression level of RdPrip was lower in the spinosad-resistant strain than in the susceptible one, and the resistant strain produced fewer progeny than the susceptible strain did. These studies support the functional role of RdPrip in female reproduction. The absence of significant mortality reduction in the R. dominica exposed to spinosad after RdPrip RNAi suggests that other aquaporins that were not knocked down may exist for the excretion of metabolized pesticides.
RESUMO
The lesser grain borer, Rhyzopertha dominica, which is a primary pest of stored products, breaks up whole grains and makes them susceptible to secondary infestation by other pests. Insecticide application is the main control measure against this borer. A resistant strain of R. dominica against the insecticide, spinosad, was selected in the laboratory. The full-length cDNA of the target site of spinosad, nicotinic acetylcholine receptor subunit α6, from R. dominica (Rdα6) was cloned and analyzed using reverse transcription PCR and rapid amplification of cDNA ends. The complete 2133-bp cDNA contains the open reading frame of 1497â¯bp encoding a 498-amino-acid protein. There are four predicted transmembrane (TM) regions, and six extracellular ligand-binding sites at the N-terminus, upstream from the first TM in Rdα6. Three mutations have been found in the resistant strain compared with the susceptible one: (1) a 181-bp fragment truncated at the N-terminus, resulting in the appearance of a premature stop codon, (2) one missing bp at the position 997, causing a frame-shift mutation, and (3) an 87-bp fragment truncated in the TM2 region. In addition, real-time quantitative PCR was applied to detect the transcriptional expression of Rdα6 in both the susceptible and resistant strains. The results indicated that the expression of Rdα6 was significantly lower in then resistant strain than in susceptible one. In conclusion, mutation of Rdα6 may cause R. dominica resistant to spinosad due to target site insensitivity.