Sequencing and characterization of the citrus weevil, Diaprepes abbreviatus, trypsin cDNA. Effect of Aedes trypsin modulating oostatic factor on trypsin biosynthesis.

Trypsin mRNA from the citrus weevil, Diaprepes abbreviatus, was reverse transcribed and amplified by PCR. A cDNA species of 513 bp was cloned and sequenced. The 3' and 5' ends of the gene (262 bp and 237 bp, respectively) were amplified by rapid amplification of cDNA ends, cloned and sequenced. The deduced sequence of the trypsin cDNA (860 bp) encodes for 250 amino acids including 11 amino acids of activation and signal peptides and exhibited 16.8% identity to trypsin genes of selected Lepidoptera and Diptera. A three-dimensional model of Diaprepes trypsin contained two domains of beta-barrel sheets as has been found in Drosophila and Neobellieria. The catalytic active site is composed of the canonical triad of His41, Asp92 and Ser185 and a specificity pocket occupied by Asp179 with maximal activity at pH 10.4. Southern blot analysis indicated that at least two copies of the gene are encoded by Diaprepes midgut. Northern blot analysis detected a single RNA band below 1.35 kb at different larval ages (28-100 days old). The message increased with age and was most abundant at 100 days. Trypsin activity, on the other hand, reached a peak at 50 days and fell rapidly afterwards indicating that the trypsin message is probably regulated translationally. Feeding of soybean trypsin inhibitor and Aedes aegypti trypsin modulating oostatic factor affected trypsin activity and trypsin biosynthesis, respectively. These results indicate that Diaprepes regulates trypsin biosynthesis with a trypsin modulating oostatic factor-like signal.

Natural Field Spread of Mild and Severe Isolates of Citrus Tristeza Virus in Florida.

The effectiveness of five different aphid control regimes in delaying movement of mild and severe isolates of citrus tristeza virus (CTV) into a CTV-free sweet orange scion on sour orange rootstock block was monitored annually for 5 years, using severe isolate-specific and isolate-nonspecific monoclonal antibodies. The average percentage of trees infected with a severe isolate of CTV was 32, 32, 20, 25, and 28 for trees treated with Temik, Temik + Meta-Systox R (MSR), MSR, stylet oil, or no aphid control, respectively, at the conclusion of the experiment (5 years). These percentages were not significantly different (P ≤ 0.05). The average percentage of trees infected only with mild isolates was 13, 10, 7, 7, and 17 for the above treatments, respectively. The infection with mild isolates was significantly greater (P ≤ 0.05) with no aphid control than with stylet oil or MSR treatments.