Assessment of temporal changes in haem and protein levels in Ixodid ticks following blood engorgement, and their use as age-grading parameters
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Nymphs of Rhipicephalus appendiculatus and Hyalomma anatolicum anatolicum were fed on rabbits and maintained in the laboratory to allow moulting and further development. At specified time intervals from 0 to 500 days after engorgement, samples of ticks were ground individually in distilled water within the wells of an agglutination plate. A 0.1 ml aliquot was removed from each and levels of haemin and protein assessed from optical density values at specific wavelengths in a spectrophotometer. Both protein and haemin levels showed an initial rapid decrease after engorgement; values did not fall to zero in either case but showed marked fluctuation throughout the study period. These fluctuations combined with the high standard errors of the results, made assessments of the physiological age of individual ticks impossible. Such fluctuating values, however, suggested the possibility that haem biosynthesis might be taking place, and this was tested by injecting the radioactively-labelled haem precursor [4-14C]δ-amino laevulinic acid into engorged nymphs, immediately following their detachment. Both tick species revealed an incorporation of this compound into their haematin content, although neither incorporated the non-haem precursor [1-14C]2-amino isobutyric acid. These results indicate an ability of ticks to synthesize haem in vivo, although the underlying reasons for such a mechanism remain unknown.