Recognition of Anaplasma marginale Theiler in Dermacentor andersoni Stiles (=D. venustus Marx) by the fluorescent antibody method. I. Smears of nymphal organs.

Smears of nymphal organs from normal Rocky Mountain wood ticks, Dermacentor andersoni Stiles, and from similar ticks that had fed on a calf infected with Anaplasma marginale Theiler were subjected to the fluorescent antibody technique. The results confirmed previous reports that anaplasmata are identifiable by this technique in smears prepared from gut contents of nymphs fed on calves infected with A. marginale on the day of detachment and for 24 hr thereafter. However, they did not confirm reports that anaplasmata can be recognized in the Malpighian tubules of infected nymphs for as much as 5 days after detachment or that multiplication by binary fission occurs in the Malpighian tubules. The nonspecific fluorescence in the gut, Malpighian tubules, and salivary glands; the intense nonspecific reactions in connective tissue and developing adult tissue; the possibility of crosscontamination (before preparation of the slide); and the possibility of cross-reactions of other microorganisms all made it impossible to identify anaplasmata conclusively, once erythrocytes were no longer available as markers. The use of fluorescent antibody methods to detect Anaplasma marginale Theiler within organs of ticks fed on infected bovines has been reported by two groups of investigators. Anthony et al. (1964) studied anaplasmata in adult female Dermacentor andersoni Stiles (= D. venustus Marx) that had partially engorged on acutely infected calves and concluded that they could be recognized in smears of the gut and excreta for 24 hr after the ticks were removed from an infected calf; however, they were unable to make critical interpretations of smears of salivary glands and reproductive organs because of the presence of nonspecific fluorescence. Friedhoff and Ristic (1966) investigated anaplasmata in nymphs of D. andersoni that had fed to repletion on an acutely infected calf and reported that anaplasmata could be detected in smears of the gut contents for 2 days and in smears of Malpighian tubules for 5 days after normal detachment; they also suggested that anaplasmata may multiply in the Malpighian tubules of the nymphs by binary fission. This suggestion had particular significance since multiplication of A. marginale within an arthropod vector had never been previously reported, nor had other authors been able to identify anaplasmata clearly within any organs of the Received for publication 17 June 1969. vector except the gut contents. Regendanz (1933) had reported identifying A. marginale in the salivary glands of infected female Boo philus microplus (Canestrini) by using classical histological techniques, but neither Cowdry and Rees (1935) with D. andersoni and D. variabilis (Say) nor Anthony (pers. comm.) with D. andersoni and D. occidentalis Marx were able to confirm these observations. MATERIALS AND METHODS