a bacterial agent identified in culture, the need remained to do studies in human subjects. The use of experimental animals was to come later (37, 38, 39). Finlay had already used human subjects in his earlier studies (14). Still, the investigators recognized the ethical implications of the studies in humans and offered themselves first. In James Carroll’s words, “Then arose the question of the tremendous responsibility involved in the use of human beings for experimental purposes. It was concluded that the results, if positive, would be sufficient justification of the undertaking. It was suggested that we subject ourselves to the same risk, and this suggestion was accepted by Dr. Reed and Dr. Lazear” (7). Carroll became the first experimental subject, accepting the risks ahead of other volunteers.
The circumstances to further study yellow fever were propitious. Following the Spanish-American War, yellow fever appeared yet again in Cuba, placing the populace and American troops at risk. Soon after arrival in Cuba, Reed and his Commission colleague, Agramonte (Fig. 7), visited an army barracks at Pinar del Rio where an outbreak was occurring. Observations made on that visit “. . . did not tend to strengthen one’s belief in the theory of the propagation of yellow fever by fomites” (30). A curious story was told of that visit. Only one of nine prisoners, well guarded in jail, had come down with yellow fever. Speculation was raised that an insect such as a mosquito had bitten the one prisoner. That speculation was buttressed by the observations of Carter of the interval between infecting and secondary cases (9, 21). It was decided to test Finlay’s theory of mosquito transmission of yellow fever.
Figure 7 The Yellow Fever Commission consisted of (upper left) Walter Reed, who led the Commission; (lower left) James Carroll, who performed the filtration experiment; (upper right) Aristides Agramonte; and (lower right) Jesse W. Lazear, who became infected in the course of the experiments and died. In a remarkably brief period of time at the turn of the 20th century, the Commission under Reed demonstrated that the disease was transmitted by mosquitoes and that it could be transmitted by filtered blood and thus was caused by a virus. (Courtesy of the Historical Collections & Services, Claude Moore Health Sciences Library, University of Virginia, except for the image of Walter Reed, courtesy of The National Library of Medicine.)
doi:10.1128/9781555818586.ch1.f7
In Reed’s words, “. . . the search for the specific agent of yellow fever while not abandoned, should be given secondary consideration, until we had first definitely learned something about the way or ways in which the disease was propagated from the sick to the well” (30). In preliminary experiments by Lazear, mosquito eggs were supplied by Finlay, and mosquitoes were raised in the laboratory, allowed to feed on yellow fever patients, and allowed to bite human subjects. First among the subjects was Carroll, who fell ill and almost perished (21). Lazear, apparently bitten by a stray mosquito in 1900, was a victim of their research efforts: he contracted yellow fever and died. The results of the experiments showed that 2 of 11 experimentally infected subjects developed yellow fever. It was concluded that “The mosquito acts as the intermediate host for the parasite of yellow fever, and it is highly probable that the disease is only propagated through the bite of this insect” (italics in the original) (29).
There followed the construction of two small buildings in an open field to compare the transmission of yellow fever by fomites with transmission by the bites of infected mosquitoes or inoculation of infected blood. The “Infected Mosquito Building” was well ventilated and divided into two compartments by a screen. The “Infected Clothing and Bedding Building” was purposely not well ventilated so as to retain any noxious effects of bed clothing, pajamas, and other items from previously infected cases. After some early discouraging results, John R. Kissinger, a soldier who Reed praised for having volunteered “solely in the interest of humanity and the cause of science” and who would accept no payment, came down with experimental yellow fever from the bites of infected mosquitoes (30). In these experiments, six of seven “non-immunes” bitten by infected mosquitoes in the Infected Mosquito Building became ill with yellow fever (32). None of the seven subjects in the Infected Clothing and Bedding Building exposed to fomites from cases of yellow fever became ill, nor did subjects become ill who had remained behind the screen, not bitten by mosquitoes.
The clarity of the design of comparison groups and the results were decisive: 85.71% infected by mosquitoes versus 0% by fomites. In the definitive publication in JAMA, “The Etiology of Yellow Fever: an Additional Note,” Reed, Carroll, and Agramonte ended with several major conclusions. In addition to confirming that “C. fasciatus serves as the intermediate host,” they determined that 12 days or more was required after contamination for the mosquito to transmit the infection. Thus, they determined experimentally what Carter had observed epidemiologically. They found that yellow fever could be transmitted by blood subcutaneously inoculated when taken from a patient on the first 2 days of the illness. They concluded that yellow fever resulting from a mosquito bite “confers immunity” against attempted reinfection with infected blood (32).
In memory of Lazear, the experimental station established by Reed, where the crucial studies were conducted demonstrating the transmission of yellow fever by mosquitoes and not by fomites, was christened Camp Lazear. Ironically, although Carroll recovered from acute yellow fever infection, he tragically died 7 years later of myocarditis attributed to that attack of yellow fever.
An important piece of the puzzle still remained to fall in place. Walter Reed and his colleagues’ final conclusion of their JAMA report was that “. . . the specific cause of this disease remains to be discovered” (32). Having turned away from that goal in their transmission studies, Carroll returned to the project. Initially confronted with local objections to further experimentation, Carroll resumed his studies in September 1901 in Cuba on the nature of the infecting agent (7). In the crucial experiment, six individuals were exposed to the bites of infected mosquitoes (33). Four did not develop yellow fever, but two did. Blood was taken from patients I and II for further transmission study, but due to an accident to the vacuum pump, the blood from patient I could not be used. The blood from patient II was divided into three aliquots of partially defibrinated and diluted serum. The first aliquot, a positive control, was left untreated and successfully transmitted yellow fever to patient III. The second aliquot was heated to 55°C for 10 minutes and failed to transmit disease to patients IV, V, and VI. Based on previous work on heat stability with toxins, Reed and Carroll argued against a toxin. The third aliquot was “slowly filtered through a new Berkefeld laboratory-filter” and the filtrate was inoculated into patients VII, VIII, and IX. Patients VII and VIII developed “unmistakable” attacks of yellow fever; patient IX remained well. The scientific data
