The Salivary Adenosine Deaminase Activity of the Mosquitoes Culex quinquefasciatus and Aedes aegypti
José M. C. Ribeiro*,
Rosane Charlab and
Jesus G. Valenzuela
Medical Entomology Section, Laboratory of Parasitic Diseases, NIAID, National Institutes of Health, 4 Center Drive, Room 4/126, Bethesda, MD 20892-0425, USA
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Fig. 1. Salivary Culex quinquefasciatus adenosine deaminase cDNA and protein sequences. The underlined sequence refers to the signal peptide identified by the program SignalP (Nielsen et al., 1997). Nucleotides are in lower case and amino acid residues in capital letters.
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Fig. 2. Adenosine deaminase activity of salivary homogenates of adult female Culex quinquefasciatus mosquitoes. (A) A cuvette containing 100µmoll-1 adenosine in 100µl of HS buffer, pH7.2, was scanned at 10min intervals following addition of salivary homogenate equivalent to 0.2 pairs of salivary glands. The arrows indicate the direction of change of the spectrum over time. (B) Differential spectra of the data in A obtained by subtracting each scan from the scan at time zero.
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Fig. 3. Adenosine (ADO) and AMP deaminase (AMP) activities in Culex quinquefasciatus salivary homogenates from adult female mosquitoes. (A) Differential spectra obtained following addition of homogenate from one pair of salivary glands to 100µmoll-1 adenosine in 100µl of HS buffer, pH7.2. (B) Differential spectra obtained following addition of homogenate from one pair of salivary glands to 100µmoll-1 AMP in 100µl of HS buffer, pH7.2. The numbers in the figure indicate the time, in minutes, that the scan was performed following the addition of the salivary homogenate.
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Fig. 5. Adenosine deaminase (ADA) activity in salivary homogenates of adult female Aedes aegypti. (A) A cuvette containing 100µmoll-1 adenosine in 100µl of HS buffer, pH7.2, was scanned at 6min intervals following addition of salivary homogenate equivalent to 0.02 of a pair of salivary glands. The arrows indicate the direction of change of the spectra over time. (B) Differential spectra for the data in A.
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Fig. 6. Salivary adenosine deaminase (ADA) activity in blood-fed and unfed female mosquitoes and in male mosquitoes. The columns and error bars represent the mean + S.E.M. of 12 determinations in individual pairs of homogenized salivary glands from Aedes aegypti and Culex quinquefasciatus.
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Fig. 7. Adenosine deaminase (ADA) activity left by adult female Aedes aegypti while probing an artificial feeder containing 50µmoll-1 adenosine in HS. (A) Continuous line, spectrum of a control solution exposed in a feeder for 20min; dotted line, spectrum of a solution after exposure to approximately 100 female mosquitoes for 20min. (B) Mosquito-exposed feeder solution (5µl) was diluted with 100µmoll-1 adenosine in 95µl of HS, and the resulting solution was scanned at 12min intervals. The arrows indicate the change in the spectrum over time. (C) Differential spectra of the data in B, obtained by subtracting each scan at 12min intervals from the scan at time zero. (D) Rate of change of absorbance at 265nm, used to calculate the specific activity of ADA.
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Fig. 8. Adenosine deaminase (ADA) and AMP deaminase (AMP) activities in serotonin-induced saliva from adult female Culex quinquefasciatus and Aedes aegypti. Individual samples of serotonin-induced mosquito saliva collected in mineral oil were transferred to 10µl of Hepes saline (10mmoll-1Hepes, pH7.2, 100mmoll-1 NaCl) and centrifuged. Enzyme levels were determined by measuring the decrease in absorbance at 265nm in microcuvettes containing 71µl of Hepes saline, 0.1mmoll-1 substrate and 4µl of the diluted saliva sample. Each salivary sample was used for one adenosine and one AMP deaminase reaction. Values are the mean + S.E.M. of five samples from each mosquito species.
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© The Company of Biologists Ltd 2001
) and AMP deaminase (
) activities. The inset indicates the elution of molecular mass (Mr) standard markers; the arrow points to the elution time of the ADA/AMP deaminase activities.