CYCLES OF PLANT VIRUSES IN INSECT VECTORS 169 



same as in the leafhopper, or in tlie same order as those in the leafhoppers. If, 

 on the other hand, the incubation periods in the vectors represented, not only- 

 such passage from gut to saliva, but also periods of virus multiplication, then 

 a rapidly multiplying virus could be expected to have a short incubation 

 period in both vector and plant, and a slowly multiplying virus would be 

 expected to have a long incubation period in both. 



In each of the years 1939 to 1942 the writer used the insect injection 

 teclmique in large-scale attempts to transmit aster yellows virus from insect 

 to insect in series. AH of these attempts were complete failures; the mortality 

 of the insects in the second passage was very high and none of the few survivors 

 was infective. At about this time a virus, clover club leaf virus, was found in 

 Agalliopsis novella (Black, 1944), and it happened to be transmitted through 

 the egg to a high percentage of the progeny (Black, 1948). It was chosen as a 

 means for obtaining proof that a plant virus could increase in its insect vector 

 (Black, 1950). A single viruliferous female weighing 1.7 mg. was mated to a 

 nonviruliferous male and was allowed to oviposit its eggs in immune alfalfa 

 plants. A portion of the progeny was tested on susceptible crimson clover to 

 determine the proportion that was infective, and the remainder of the 

 progeny were reared throughout their lives on alfalfa. From the proportion 

 of insects that infected crimson clover the number of infective insects among 

 all the progeny of the female was estimated; the reciprocal of this number 

 constituted a conservative measure of the average dilution of the starting 

 virus in each of the progeny. 



Females among the progeny kept on alfalfa all their lives were mated with 

 nonviruliferous males and the experiment continued in such a way that 

 selection of the main hne of descent was independent of virus concentration. 

 After continuing the experiment for more than 5 years through 21 genera- 

 tions, it was calculated that the dilution of virus in the original female was 

 equivalent to at least 1/10-*', whereas the maximum number of virus particles 

 that she could have contained was estimated to be 10^^. The virus was also 

 passed by graftage from one Vinca rosea plant to another through 30 passages 

 in series. The experiment provided proof that this virus multiplied in both 

 insect vector and plant host. 



While the third edition of Bawden's book was in press he was sho\Mi these 

 data when visiting the United States. As a result, he included an addendum 

 in the 1950 edition agreeing with the evidence for the abUity of this virus 

 to maintain itself indefinitely in its insect vector. 



Maramorosch (1952a), using the insect injection technique, succeeded in 

 the serial transmission of aster yeUows virus from leafhopper to leafhopper 

 under conditions which assured no replenishment of virus from plants. The 

 insects were maintained on immmie plants or left on susceptible plants for 

 only very short periods. At each transfer he inoculated the insects with a 



