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United States Department of Agriculture

Agricultural Research Service

Research Project: IMPROVEMENT OF PEST RESISTANCE AND QUALITY TRAITS OF SOYBEAN Title: Characterization of a chitin synthase encoding gene and effect of diflubenzuron in soybean aphid, Aphis glycines

Authors
item Bansal, Raman -
item Mian, Rouf
item Mittapalli, Omprakash -
item Michell, Andrew -

Submitted to: International Journal of Biological Sciences
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: September 10, 2012
Publication Date: October 25, 2012
Repository URL: http://handle.nal.usda.gov/10113/58206
Citation: Bansal, R., Mian, R.M., Mittapalli, O., Michell, A.P. 2012. Characterization of a chitin synthase encoding gene and effect of diflubenzuron in soybean aphid, Aphis glycines. International Journal of Biological Sciences. 8(10):1323-1334.

Interpretive Summary: Soybean aphid, A. glycines, is the number one insect pest of soybean in USA and controlling this insect costs hundreds of millions of dollars every year. Chitin synthases are critical enzymes for synthesis of chitin and thus for subsequent growth and development in insects. We have identified and characterized a chitin synthase gene (CHS) from cDNA of the soybean aphid. The full-length cDNA of CHS in A. glycines (AyCHS) was 5802 bp long with an open reading frame of 4704 bp that encoded for a 1567 amino acid residues protein. Real-time PCR analysis revealed that AyCHS was expressed in all major tissues (gut, fat body and integument), however, it had the highest expression in integument (~3.5 fold compared to gut). Interestingly, the expression of AyCHS in developing embryos was nearly 7 fold higher compared to adult integument, which probably is a reflection of embryonic molts in insects. Expression analysis in different developmental stages of aphid showed that AyCHS was expressed in all growth stages but had the highest expression in second instar stage. Further, through leaf dip bioassays, we tested the effect of diflubenzuron, a chitin-synthesis inhibitor, on aphid survival, fecundity and body weight. When fed with soybean leaves previously dipped in 50 ppm diflubenzuron solution, aphid nymphs suffered significantly higher mortality as compared to control. Aphid nymphs feeding on diflubenzuron treated leaves also showed significantly enhanced expression of AyCHS compared to nymphs on untreated leaves. The current study provides a platform to develop novel strategies to manage this pest using chitin-synthesis inhibitors and using molecular technology for knocking down AyCHS expression.

Technical Abstract: Chitin synthases are critical enzymes for synthesis of chitin and thus for subsequent growth and development in insects. We have identified and characterized a chitin synthase gene (CHS) from cDNA of Aphis glycines, the soybean aphid, a serious pest of soybean. The full-length cDNA of CHS in A. glycines (AyCHS) was 5802 bp long with an open reading frame of 4704 bp that encoded for a 1567 amino acid residues protein. The AyCHS protein had a molecular mass of 180.05 kDa and its amino acid sequence contained all the signature motifs (EDR, QRRRW and TWGTR) of chitin synthases. Real-time PCR analysis revealed that AyCHS was expressed in all major tissues (gut, fat body and integument), however, it had the highest expression in integument (~3.5 fold compared to gut). Interestingly, the expression of AyCHS in developing embryos was nearly 7 fold higher compared to adult integument, which probably is a reflection of embryonic molts in hemimetabolus insects. Expression analysis in different developmental stages of A. glycines revealed that AyCHS was expressed in all stages but had the highest expression in second instar stage. Further, through leaf dip bioassays, we tested the effect of diflubenzuron, a chitin-synthesis inhibitor, on A. glycines’ survival, fecundity and body weight. When fed with soybean leaves previously dipped in 50 ppm diflubenzuron solution, A. glycines nymphs suffered significantly higher mortality as compared to control. A. glycines nymphs feeding on diflubenzuron treated leaves also showed significantly enhanced expression (~1.7 fold) of AyCHS compared to nymphs on untreated leaves. The current study provides a platform to develop novel strategies to manage this pest using chitin-synthesis inhibitors and using RNAi by knocking down AyCHS expression.

Last Modified: 11/23/2014
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