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PURCHASED FROM THE INCOME OF THE

SAMUEL WHEELER WYMAN

LES PRIX NOBEL 1902

ABSTRACT

RESEARCHES ON MALARIA

BEING

THE NOBEL MEDICAL PRIZE LECTURE FOR 1902

BY

RONALD ROSS

FELLOW OF THE ROVAL COLLEGE OF SCRGEONS

FELLOW OF THE ROVAL SOCIETY

COMPANION OF THE BATH

PROFESSOR OF TROPICAL MEDICINE, UNIVERSITY OF LIVERPOOL

STOCKHOLM

Kt.-NGL. BOKTRYCKERIET. P. A. NORSTEDT & SdNER
190^

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in 2011 with funding from

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RESEARCHES ON MALARIA

BEING

THE NOBEL MEDICAL PRIZE LECTURE FOR 1902

BY

RONALD ROSS

FELLOW OF THE ROYAL COLLEGE OF SURGEONS

FELLOW OF THE ROYAL SOCIETY

COMPANION OF THE BATH

PROFESSOR OF TROPICAL MEDICINE, UNIVERSITY OF LIVERPOOL

STOCKHOLM

KUNGL. BOKTRYCKERIET. P. A. NORSTEDT & SONER
1904

/^Yi^

Y6

RESEARCHES ON MALARIA.

Contents.

Page

1. Preliminary ■ I

2. The Discovery of the Parasite of Malaria 3

3. The Problem of the Mode of Infection 4

4. First Researches in India: 1S89 — 94 6

5. Return to England: 1894 7

6. The Theories of King, Laveran, Koch, and Manson 8

7. Nature of Proposed Investigations 12

8. Preliminary Observations at Secunderabad; 1895 13

9. Secunderabad; 1895. The Motile Filaments in Mosquitoes 14

10. Difficulty of the Task. New Methods Devised 15

11. Bangalore: 1895 — 97- Progress of Work 19

12. The Sigur Ghat: 1897 25

13. Secunderabad; 1897. The Fundamental Discovery 29

14. Interruption; September 1897 — February 1898 39

15. Calcutta; February — April 1898. The Theory Proved 42

16. The .Darjeeling Terai: April — June 1898. Efforts to Obtain Assistance 49

17. Calcutta; June — August 1898. The Route of Infection > 53

18. Darjeeling District; August — September 1898. Kala-Dukh 60

19. Assam; September — November 1898. Kala-Azar 61

20. Calcutta; November 1898 — February 1899. The Work Confirmed 63

21. England; March— July 1899. Foundation of the Liverpool School of Tropical

Medicine 67

22. Sierra Leone: x\ugust — September 1899. The Investigation Completed 70

23. Confirmation and Extensions 73

i) The Work of Koch 73

2) The Italian Writings 75

3) The Commission of the Royal Society 82

References. Description of Plates. Plates 84 et seq

RESEA:RC^iE5^^^^%|lALARIA

(Being the Nobel AIedicalv.£ri4e/ttec!Tli9^^|?trijbe 1902, delivered at Stockholm

onTRe laflTDecember.)*

Preliminary. — Malarial Fever, or, as it is often called, Paludism or
Tntermittent Fever, is perhaps the most important of all diseases which
afflict humanity. Broadly speaking it is spread over almost the whole of
the tropics, and also extends into many countries which possess temperate
climates — being found as far north as Sweden and Canada. Although,
happily, it is not a very fatal disease, yet it is generally so prevalent in
the countries in which it exists that the sum of the illness which it causes
is immense. To take for instance the great country of India with its
enormous population of nearly three hundred millions, we find from the sani-
tary returns of the government that the deaths from fever alone are given at
4,919,591 for the single year 1900; and average roughly about five million
■deaths yearly — a population nearly as large as that of Sweden and Nor-
way. Although it is not possible to state that all this fever is malarial
fever, there are reasons for thinking that most of it must be such. From
the more exact returns of the army and of the jail prisoners in India —
returns attested by medical men — we find that in 1900, out of the total
of 305j927 persons, no less than 102,640 were admitted into hospital for
malarial fever during the year; and even this large figure is below the
truth, because in India many slight cases of fever are not admitted into
hospital at all. The following table taken from the returns of the British
Troops in India for 1900 will enable us to compare the sickness due to
malaria and to other diseases respectively:

* It should be understood that the lecture as gi^-en on this occasion was only an abstract
of the present publication.

Researches on Malaria. I

Average Strength of Troops 6o^6^j.

Disease Admissions

Malarial Fever .... 19,445

Enteric Fever 970

Other Fevers 1.479

Dysentery 1,561

Hepadc Congestion . . i.oio

Hepatic Abscess .... 156

Heat Stroke 174

Cholera 107

Contagious Diseases . . 18,049

leaths

Cons

tantly sick

50

710

290

141

2

67

52

108

5

68

95

15

52

8

89

2

14

I

,650

Total 42,951 649 2,769

It should be noted that the death rate for malaria is here far below
the truth; because, the disease being often very chronic, many of the
Avorst cases are invalided to Europe; while in others death is often recorded
as being due to intercurrent affections, such as pneumonia or dysentery,
even though malaria may have been the original or principal cause of the
fatal result.

Similar statistics will be found in most of the tropical countries of
the world where statistics are kept at all. Even in such a temperate climate
as Italy, the annual number of cases amounts, according to Celli, to some-
thing like two millions, while the number of deaths may be fifteen thousand
a year. For the great continent of Africa we have, of course, no figures ^
but we know from the important discovery of Koch, confirmed by many
German and British workers, that between fifty and a hundred per cent
of the negro children always remain infected — from which also we may
assume that the terrible infantile mortality among negroes is largely due
to this disease.

But malarial fever is important, not only because of the misery which
it inflicts on mankind, but because of the serious opposition which it has-
always given to the march of civilization in the tropics. Unlike many
diseases, it is essentially an endemic, a local, malady; and one which un-
fortunately haunts more especially the fertile, well-watered and luxuriant
tracts — precisely those which are of the greatest value to man. There
it strikes down, not only the indigenous barbaric population, but, with
still greater certainty, the pioneers of civilization, the planter, the trader,
the missionary, and the soldier. It is therefore the principal and gigantic
ally of barbarism. No wild deserts, no savage races, no geographical dif-

ficulties have proved so inimical to civilization as this disease. We may
almost say that it has withheld an entire continent from humanity — the
immense and fertile tracts of Africa; what we call the dark continent
should be called the malarious continent; and for centuries the successive
waves of civiHzation, which have flooded and fertilised Asia, Europe, and
America, have broken themselves in vain upon its deadly shores.

2. The Discovery of the Parasite of Malaria. — From the first
then, the study of this potent foe of mankind has given a great occupation
to science. It is not within my province at this moment to detail the
early steps by which science gradually penetrated the mystery — steps,
however, which are not the less interesting to follow. Though it was well
known to the ancients, the disease was not clearly differentiated from other
fevers until much later. Towards the middle of the seventeenth century,
however, physicians recognised that in cinchona bark we possess a drug
which is a specific for a certain class of fevers, namely the intermittent
fevers. As Kelsch and Kiener remark, this discovery was not only an
immense therapeutical benefit, but also led to a notable pathological ad-
vance, because it enabled Morton and Torti to prescribe the exact limits
of the disease curable by the medicine, namely malarial fever; and the
works of these writers, especially Torti, who without possessing thermo-
meter or microscope accurately described the intricate course of the dis-
ease, are among the most admirable works of medical science. At the
end of the seventeenth and the beginning of the eighteenth century Morton
and Lancisi elaborated another important conception, that the disease is
produced by some poison which enters the body from without ; and the latter
especially clearly understood what may be called the great law of malarial
fever — that it is connected with stagnant water on the ground. The
next great advance was made in the middle of last century by Meckel,
Virchow, Planer, Arnstein, Frerichs, and others, who discovered that the
disease is characterised by the presence in the blood and some tissues of
a peculiar black granular substance, the malarial pigment or melanin; and
this observation led directly to the great discovery of Laveran in 1880,
that the melanin is produced within multitudes of minute amoeboid
parasites which live within the blood corpuscles of the patient — a dis-
covery which not only illuminated the whole subject of malaria, but, by
opening a new department of parasitic pathology, has put the name of
Laveran in the place of honour beside those of Pasteur, Lister, and Koch.

The work of Laveran and of those who followed him affords one of
the most beautiful and useful chapters in the whole book of science; and
I wish that it were possible to deal with it here at length. We owe to
Danilewsky, Theobald Smith and others the discovery of similar para-
sites in the blood of many vertebrates, and to Laveran and Golgi the
determination of several important laws concerning the whole group of
these organisms. Marchiafava, Celli, Mannaberg, Metchnikoff, Canalis,
Antolisei and many others added important details; Kelsch and Bignami
made minute clinical and pathological studies; Romanowsky discovered the
best method of staining the parasites; Gerhardt and others produced in-
fection by inoculating the blood of patients into healthy persons; and
Richard, Councilman, Vandyke Carter, Osier, Plehn and numerous other
skilled observers confirmed those results in many parts of the world.
The principal conclusions reached by this mass of investigations are as
follows: —

(i). That Laveran's parasite is the cause of malarial fever.

(2). That it is a sporozoon belonging to a group probably allied to
the Coccidiidae, of which other members are found in birds; and that
somewhat similar but more distantly related hsemocytozoa are found in
other vertebrates.

(3). That the organisms propagate in the blood by spore formation.

(4). That there are probably at least three varieties of the human
parasites, which cause respectively the quartan, the tertian, and the irregular
(pernicious or aestivo-autumnal) fevers.

(5). That the paroxysm of fever commences with the release of the
spores.

(6). That with all varieties of the parasites, there are certain forms
which do not produce spores, but which, shortly after blood containing
them is drawn from the host, emit certain singular motile filaments; and
that the nature and functions of these forms still required further investigation.

3. The Problem of the Mode of Infection. — But even after all
these fine discoveries, there still remained for solution a problem of the
greatest difficulty and of the greatest importance. We had discovered
the pathogenetic organisms of malarial fever, and had studied them and
their effect with the greatest care. This was much, but not all; it suf-
ficed for the treatment of individual cases; but not for the prevention or
extirpation of the disease on a large scale. For this we were obliged to

5 -

seek a wider knowledge: the parasites occur in the human blood — but
how do they arrive there? On this scientific question turned the whole
prophylaxis of malaria — a subject the importance of which in connec-
tion with the future development of many of the richest portions of the
world's surface I need not enlarge upon. Ignorant of the route of entry,
we could rest our prophylaxis only upon an unsatisfactory empirical basis;
cognisant of it, we might hope to stamp out the plague even in its most
redoubtable haunts. It is my privilege in this lecture to describe par-
ticularly the steps by which this great problem has at length received its
full solution.

In what manner precisely does the malarial infection reach the human
blood? From early times certain cardinal facts regarding the disease have
been known to us and have limited the area of investigation regarding
this question. It has been recognised, first that malarial fever is essentially
an endemic disease — that is, that it does not easily spread from man to
man independently of locality, as do for instance, small-pox or plague;
secondly that it adheres especially to warm localities where there is much
stagnant water, such as marshes. Upon these facts, themselves perfectly
true, numerous hypotheses have been constructed; notably the one, dating
from the times of Lancisi and Morton, that the disease is due to miasmata
exhaled from the stagnant water — whence indeed the word malaria has
originated; and later the allied theory of the telluric miasm, according to
which the soil possesses a poisonous effluent so powerful at certain spots
that it can there produce fever in man. It was even thought that when
the surface of the ground is disturbed, this effluent escapes like a gas, in-
fecting all those who live in the vicinity. These speculations afi"ord an
interesting example of the manner in which the human mind is apt to
embroider fact with hypothesis. It is a fact that malarial fever is connected
with stagnant water; but that the connection is due to an aerial emanation
from the stagnant water was only an hypothesis which has never received
experimental verification. Nevertheless it was almost universally accepted
until the true explanation of the connection referred to was given in the
manner which I shall presently describe.

The discovery of the pathogenetic organism by Laveran produced
but little change in our ideas on this point. It was simply thought that
the parasites must be capable of saprophytic life in stagnant water, and
may enter the body by the inhalation of watery vapour or by infected
drinking water; and, indeed, efforts to obtain experimental proof of these

conceptions were quickly made, especially by Calandruccio, Marino, Agenore,
and Celli [8, 7, 4], who endeavoured to infect healthy persons by means
of water brought from notoriously unhealthy sources. The experiments
proved, however, entirely negative — somewhat to the surprise of those
who were acquainted with them. At the same time parallel enquiries
were commenced to ascertain the saprophytic stage of the parasites; and
Grassi and Feletti found an amoeba {Auia^ba gutttdd) which they thought
might be the parasites in their free condition [10]. Their work recalls that
of Crudeli and Klebs who, before Laveran's discovery, claimed to have
found the cause of malaria in the form of a bacillus — which they asserted
abounds in the water and soil of malarious localities, especially in the
lowest stratum of the air, and gives typical intermittent fever to rabbits and
other animals. All these observations are now proved to have been unsound.

4. First Researches in India; 18 8g — g^. — It is, I understand, the
principal duty of those who are called to the high honour of presenting
the lecture of the Nobel Medical Prize to give in it an account of their
own researches; and I shall therefore begin my personal narrative at
this point. * I had entered the medical service of the government of India
in the year 1881; but, although many opportunities for studying the ma-
larial problem had been given me, I was not specially attracted to it until
the year 1889, when I first began to observe many facts at variance with
the telluric hypothesis which had been instilled into me during my curri-
culum. I noted especially that the disease had a much more limited and
localized prevalence than could be explained on any theory of aerial con-
vection; I found that outbreaks often appeared to occur among troops
merely as the result of chill or fatigue; and that in many instances the
symptoms accorded ill with the classical descriptions. These observations
provoked in me much dissatisfaction with accepted theories; and gradually

* More or less brief abstracts of these investigations have already been published [54.
68], but their brevity has only had the result of permitting the genesis of many errors
regarding the real nature of the task. I have therefore thought it best to give in this publication
a fuller, and indeed almost autobiographical, narrative of the successive events. It is scarcely
possible, except by this means, to present a true picture of the difficulties in the way of re-
solving this intricate problem. As so often happens in science, the most important part of
the investigations really consisted of the initial failures; and I have therefore described these
negative results in as much detail as is given to the discovery of the pigmented cells and of the
life history of Proieosoina which afterwards gave the fundamental solution of the problem. It
should be added that my work was minutely recorded, not so much in publications, as in a
long series of letters to Manson, Laveran, and Xuttall. and that extracts from these letters
are now about to be published, together with reprints of some of my papers.

led me to the task of reviewing the whole subject by close analysis. Un-
fortunately at that time it was extremely difficult to obtain in India any
■of the more recent literature on the subject; and even the discovery of
Laveran (1880) had scarcely penetrated there as yet — much less the work
of Golgi, Danilewsky, Marchiafava and Celli- I was therefore forced to
rely almost solely on my own observations and thoughts; and at first fell
into the mistaken conception, parallel with that of Broussais, that the dis-
-ease may be due to intestinal autointoxication; and I published some pa-
pers supporting this view [12 — 16]. In 1892 however several writers began
to ventilate Laveran's work, but most unfortunately described, not the
•parasites of Laveran, but a number of artifacts.* The error was speedily
■detected and exposed [17, 18, 20, 21]; but naturally led me (and many
others in India) to doubt the whole discovery. As happened with many
others, although in pursuance of these studies I had made a laborious
examination of malarial blood for some years, I had failed entirely to
find the true parasite at all.** Up to the year 1894 therefore,, my work,
though it gave me an invaluable training for what was to come, remained
in itself quite ineffective.

5. Return to England; zSg^. — In 1894 I obtained furlough to Eng-
land; and immediately on arrival sought the advice of Professor Kanthack,
He assured me that I was mistaken in doubting the truth of Laveran's
discovery, and referred me to Dr. Manson (now Sir Patrick Manson).
Manson, to whom the parasite had been previously demonstrated in Eng-
land, now in his turn showed it to me; and also made me acquainted
with the invaluable and illuminating monographs of Mannaberg, and of
Marchiafava and Bignami. I now collected my studies in the form of an
essay (unpublished) in which I discussed the position of the malarial pro-
blem at the time, and which was accorded the Parkes Memorial Prize for
1895. In November 1894, Manson communicated to me his hypothesis,
just formed by him, that the mosquito is the intermediary host of the
malaria parasite, as he had proved it to be of Filaria noctiirna. I was
immediately and powerfully struck with this hypothesis, and at once de-
termined to give it close experimental examination on my return to India.

* Vandyke Carter had accurately followed Laveran in Bombay in 1S87; but I did no
see his work until 1894.

** As I found subsequently, one reason for this was that I had been working principally
with old aestivo-autumnal infections in which the larger and more obvious parasites (crescents)
-were scarce.

At the same time I remembered that the same hypothesis had been men-
tioned by Laveran, and I told Manson of the fact. It was not until 1899^
after the solution of the problem, that Nuttall informed me of the earlier
theories of King and Koch enunciating the same view. Consequently I
have always thought it proper to state that my own work on that part
of the malarial problem which flowed from the mosquito theory was based
on the hypothesis of Manson and Laveran. But I do not wish by this
admission to underrate those of King and Koch; and I shall now enter
upon a short digression in order to examine all these very interesting:
hypotheses together.

6. TJie Theories of King^ Laveran, Koch and Manson. — As already
mentioned, when the malaria parasite was discovered everyone who remem-
bered the old telluric and miasmatic hypothesis thought that it must live
a saprophytic existence in marshes; and up to 1894 Grassi's Amwba gnt-
tula was looked upon as being possibly the free form of the organism irt
water. Another interpretation of the connection between malarial fever
and stagnant water had, however, been noted as early as 1883 in a re-
markable paper by King [2]. King advanced the view that the malarial poison
is carried from the marsh to the human being by the bites of mosquitoes-
which breed in marshes; and he gave with great dexterity no less thaa
nineteen reasons in favour of this position — reasons based entirely on
epidemiological considerations such as the frequency of infection in warm
moist climates, in the evening, in the lower storeys of houses, and so on.
He refers to a previous enunciation of this conjecture in papers by Craw-
ford in 1807 and Nott in 1847, '^'^^ apparently lost; and he quotes
Hanson's filaria-mosquito work as a strong reinforcement of his views; but
he is evidently ignorant of Laveran' s discovery, which was then slowly
fighting its way into recognition. A fuller account of his excellent paper
will be found in Nuttall's history [65, 66\.

Laveran's conjecture was first given briefly in 1884 [3, p. 457], evi-
dently independently of King. Seven years later he mooted the same
idea, still very briefly and without giving many reasons [11, p. 147]. The
similar conjecture of Koch was not published at all; but in a letter to me
he says that the mosquito theory occurred to him during his first visit
to India in the winter of 1883 — 4, and that R. Pfeiffer mentioned the mat-
ter pubHcly in 1892 (see Koch's letter in section 23).

As already stated Manson did not arrive at his hypothesis until near
the end of 1894, when he drew attention to it (after mentioning it to me)
in a short article [22]. He based it, not upon the epidemiological con-
siderations of King, but upon a very powerful parasitological argument of
his own — which was as follows. The work of Laveran, Golgi, Marchia-
fava, Celli and others had clearly established that the general Ufe-cycle of
the parasites within the vertebrate host consists of a process of schizogony
or asexual spore formation, by means of which the organisms proliferate
indefinitely in the blood. But in addition to the sporocytes existing for

Figure I. Gametocytes producing motile filaments; the tertian parasite

(I — 3): the quartian parasite (4- — 6): the sestivo-autumnal parasite

(7 — 1 2). From page 644 of Manson's paper [26].

this purpose, all observers from the time of Laveran had observed certain
large cells, which, while they were evidently mature as regards size, did
not produce spores and appeared to have no function within the body.
Laveran showed, however, that a few minutes after blood containing these
cells was withdrawn from the circulation they underwent a singular change
— that is, they gave issue to a number of long, actively motile filaments,.

lO

capable of separating themselves entirely from the parent cell, and pro-
gressing independently among the blood-corpuscles. There had already
been a long discussion about these forms. Grassi, followed by Bignami
and many other Italians, considered them to be forms of degeneration,
and held that the motile filaments were products of a kind of death agony
in vitro. The reason given for this view was that the motile filaments
contain no chromatin (which is not true); but in my opinion these observ-
ers had not considered them with sufficient attention, or they could not
have thought them to be dying bodies. On the other hand Laveran,
Danilewsky and Mannaberg, who had studied them closely, came to the
opposite conclusion that they constitute in some way the highest stage of
the parasite; and Mannaberg even conjected that they may be concerned
in the passage from the intracorporeal to the extracorporeal stage of the
organisms — '■ though he did not indicate the route by which he thought
the passage was made. Manson's speculation broke in at this point. He
accepted Mannaberg's position; and noted also the general law that para-
sites must attain some means of passing (at least by their progeny) from
the already-infected individual into a fresh individual; that the parasites
of malaria being contained within the closed cavity of the circulation
cannot escape from it except by the intervention of some external agency
(e. g. a suctorial insect); that the position as regards these parasites was
indeed the same as that of the filaria embryos which he had shown require
the intervention of a mosquito to escape from the infected host; and that
the epidemiological laws of malarial fever suggest a possible connection
with the same insect. Hence it flashed upon him that the motile filaments
mentioned above are really flagellate spores, which, when the parent cells
are ingested by the mosquito, escape and enter the insect's tissues, de-
veloping in them into some form analogous to that of the organisms in the
human blood.

Manson continued the speculation to a further point, especially in a
later publication [26]. It will be remembered by those who have studied
his works that in his original investigations on the development of filaria
embryos in mosquitoes he had failed to ascertain that the insects live for
more than a few days; he had thought that after a single meal of blood
the mosquito lays her eggs on the surface of water and dies in the act
of doing so. Consequently when he had traced the development of the
embryos to the stage which they reach in the insect's thorax he inferred that
that w^as the whole development, and that after the death of the host on the

II

surface of the water the embryos escaped into the latter and finally in-
fected man by the digestive tract.* And he now applied the analogy to
the malaria parasites, and thought that, similarly, after the insect's death,
they enter the water and infect man either by drinking-water, as he as-
sumed for the filariae, or by the old machinery of the aerial miasma. Thus
Hanson's hypothesis suggested a clue only to the departure of the para-
site from the human host; it did not attempt to define the route of entry,
the exact mode of infection. In these points he admitted that his specu-
lation was looser, and research has shown that he was wrong — certainly
with regard to the malarial parasite and probably even with regard to
filariae. In another point also he was wrong — the motile filaments are
not flagellate spores, as he thought they were. I remember mentioning
to him at the time that they might possibly be of the nature of sperms
— an idea which was suggested to me by Lewis, who conjectured that
his trypanosoma might be that of nature [6, p. 638]. We shall see later
what they really are. But these errors were immaterial. The fundamental
part of his hypothesis was the close and powerful argument to the effect
that the motile filaments and the parent cells from which they spring must
be meant to infect the mosquito in some manner. This was more than
a hypothesis; it was a great and illuminating induction. It gave the re-
quired clue to further research; and without it I am convinced the malaria
problem would not have been solved at all and we should still be en-
gaged in a laborious and hopeless search for the parasites in water and air.
Cogent as Manson's arguments appeared to me, they were far from
convincing to most other students of malaria; and in fact no one else
took the trouble to investigate the matter in spite of its immense import-
ance to humanity. In 1896 indeed, Bignami wrote a long and dexterous
article attacking the whole induction [29]. He still refused to believe that
the motile filaments were anything but the result of death in vitro, and
added that if the induction were true, malarial fever could be propagated
by patients living in the presence of mosquitoes — which he refused to
•consider possible. At the same time he evolved a theory of his own to
the effect that mosquitoes become infected with the parasites when in the
larval stage in water, and then inoculate them into man during puncture.
Thus while Manson thought mosquitoes carry the parasite from man to
the marsh, Bignami thought that they carry it from the marsh to the
man. The latter view does not appear to me a very philosophical one'

* Or possibly »by piercing the integuments*.

12

since it presupposes the possibility of organisms living normally partly a..
saprophytic and partly a parasitic existence in the mosquito, and then
being suddenly transferable on exceptional occasions to man. Bignami's
theory, however, was exactly that of King given much more forcibly thir-
teen years earlier. He mentioned that he had previously attempted to in-
fect men by gnats brought from malarious localities, but that the attempts
had failed. He also referred to experiments made by Calandruccio, who
had failed to observe any development of the parasites in the stomach of
mosquitoes fed on malarial blood. Lastly he cites another most valuable
analogy in favour of the mosquito theory of malaria, namely that of the
Pyrosoma bigeminnni^ a parasite of cattle allied to the parasites of malaria,
and known by the brilliant researches of Theobald Smith and Kilborne
to be carried by ticks [19]. Koch also had used this analogy, but I think
that both Manson and myself had overlooked it somewhat unduly.* Cu-
riously enough all this time it seems to have occurred to no one that the
mosquito may act in both roles imagined by King and Manson severally
— that it may both take the parasite from the patient and also inoculate-
it into healthy persons. I traversed Bignami's criticisms in an article
which will be referred to later (section ii).

In considering the merits of these various hypotheses we must always-
remember that all of them have found no little support from Manson's-
original discovery of the development of Filaria hancrofti in mosquitoes.

7. Nature of Proposed Investigation. — We must now return to my
own labours. As already mentioned, directly I became acquainted with?
Manson's induction, I determined to continue my investigation of the-
malaria problem entirely on this new basis.

Before my departure for India I discussed with Manson the best me-
thod of procedure.** We agreed that the proper course would be to select;

* The development of the Pyrosojna in ticks is still unknown, though the second host has-
long been recognised. It should be understood that the history of this organism, and of
the filaria in mosquitoes, while adding great force to the mosquito theory of malaria, gave us
no information regarding the form and position of the malaria parasites in mosquitoes, nor of
the species of insect concerned. The Pyrosoma is not very nearly related to the malaria parasites, and
the filaria not at all. I was not aware of the work of Smith and Kilborne until much later.

** We thought that Manson himself could not undertake the work in England, but this-
perhaps would not have been as difficult as we supposed. The parasites have now been cul-
tivated in the local Anopheles at Hamburg and have been found in them in Holland. Work
might easily have been done in this direction at home, while I was labouring in India. At
all events I should have been greatly assisted by a study of British gnats.

13

patients whose blood was rich in gametocytes (the name now given to
those forms of the parasite of which some produce motile filaments); to
allow mosquitoes to bite these patients; and to attempt to trace in the
tissues of these insects the development of the said motile filaments (which
we thought were flagellate spores). In fact it was proposed that I should
adopt exactly the procedure employed by Manson in regard to Filaria
iancrofti. It seemed necessary only to follow the motile filaments, after
their escape from the gametocytes contained in the ingested blood in the
mosquito's stomach, to their supposed destination within some kind of
cells of the insect's tissues (e. g. the stomach or blood cells) — apparenth'
an easy task. It was true we anticipated, on the analogy of the filaria,
that not all species of mosquitoes would be amenable to the malarial in-
fection, and we recognised that this doubt would increase the difficulties;
but we hoped readily to distinguish the proper species by its particular
prevalence in very malarious localities. The motile filaments being traced
to their habitat in particular cells of the insect, we thought that it would
be easy to observe their further development, and to watch their escape
into water after the host's death. This done we should be able to identify
the extra corporeal form of the parasites in water, air, or dew, and to
ascertain exactly the route of infection of man.

8. Preliminary Observations at Secunderabad i8gj. — I reached India
in 1895 and found myself appointed medical officer of a regiment of native
soldiers stationed at Secunderabad and suffering much from malarial fever.
A survey was immediately made of the malarial parasites existing among
these men and I found myself able to confirm for India, in almost every
•detail, the specialised work of the Italians and of Mannaberg.*

At the same time I examined the mosquitoes which abounded in the
barracks and hospital. Before leaving England I had made many attempts
to obtain literature on mosquitoes, especially the Indian ones, but without
success except for some brief notes in encyclopedias; and I did not even
■clearly recognise the identity of mosquitoes and gnats, but thought that

* My regiment was stationed near a small marsh and suffered badly, while another regiment
situated only a mile to leeward of the same marsh escaped. My regiment suffered from the
-sestivo-autumnal and tertian varieties of parasite, quartan being quite absent; but in the neigh-
bouring regiments of this great garrison quartan abounded — a fact which confirmed me in
favour of the view that these varieties are distinct and not interchangeable forms. The
work of Crombie and myself was the first done in India on this basis, that of Vandyke Carter
■(1887) being done on the basis of Laveran's works.

14

the former constituted a special division of the Cuhcidse.* Consequent!}^
I was forced to rely entirely on my own observations; and I noted that
the various species of mosquitoes of the locality belonged apparently to-
two different groups, separated by many traits, and called these groups
for my own convenience, brindled mosquitoes and grey mosquitoes. It
was not until 1897 that I clearly recognised a third group which I called
spotted-zvinged mosquitoes (see sections 12 and 13).

As the grey and brindled mosquitoes abounded round the infected,
barracks, it was naturally thought likely that they were concerned in the-
propagation of the disease. After some initial difficulties I caused numbers-
of them (especially the brindled mosquitoes) to be fed on persons with
the gametocytes of ^tstivo-autumnal fever (cresents) in their blood. It
should be noted that from the first I employed for this purpose only
mosquitoes bred in captivity from the larvae, and not insects caught at
random in the houses. There were two excellent reasons for this; firsts
that the insects caught at random might have already fed themselves pre-
viously and have thus, for all I knew, acquired various parasites which,
might confuse my results; and, secondly, because it is easier to obtain the
insects in numbers by collecting their larvae and keeping them in vessels
until they hatch out from the pupae, rather than by catching each sepa-
rately by hand. The mosquitoes were fed by being released from the
breeding-jar into a mosquito-net within which the patient was placed, the
gorged insects being subsequently caught in bottles and dissected as.
required.

From the first I kept careful notes of my observations, and also
recorded them in letters to Manson sent by almost every weekly mail,
except when later, being very busy at Bangalore, I was obliged to reduce
both notes and letters. The note-books and letters are still in my hands.

9. Secunderabad; iSgj. The Motile Filaments in Mosquitoes. — The
first point requiring study was the process by which the motile filaments
escape from the parent cells (gametocytes) within the stomach cavity of
the mosquitoes. The process had been frequently watched in vitro (ia
slides of liquid blood prepared for the microscope), and was known to
occur in from about 10 to 30 minutes after the blood is drawn from the
patient; but it was now necessary to follow it in the mosquito. The in-

* In spite of repeated attempts to obtain such literature I remained in the same predica-
ment until I returned to England in 1899.

15

sects were killed from one minute to several hours after feeding, the
stomach being then extracted and its contents examined in the fresh state.
I was obliged to invent the technique for myself; and my first successful
dissection was made on the 13th May. Within a few weeks I made a
fairly complete study of the subject and ascertained an encouraging fact.
In vitro, doubtless owing to the unnatural conditions, only about five per
cent of the crescents give issue to the motile filaments; but I now found
that in the mosquito's stomach something like sixty per cent of them do
so. It was also noted that the preliminary stages of the process, namely
the swelling up and rounding of the crescents, were much more constantly
seen in the blood ingested by the insect than in vitro. This was of im-
portance because it showed at least that the insect's stomach is a more
favourable l&cus for the process than an ordinary specimen of blood is.
I observed also that a considerable percentage of the crescents (about one-
third) never produced motile filaments at all,* even after the lapse of
several hours and within the insects; and I noticed that those which re-
fused to emit them had a slightly different appearance to the others. At
the time I thought that they were parasites which had been killed in some
manner during ingestion; but when I repeated the experiments next year
I saw cause to doubt this, and felt some difficulty in explaining why all
the crescents did not emit filaments as they should have done according
to Manson's hypothesis regarding their nature.

A description of these first results was written in June, but was not
published until the end of the year [24].

10. Difficulty of the Task. Nezv Methods Devised. — The fact, then,
was established that the gametocytes are not immediately killed in the
mosquito's stomach (as might well have happened), but indeed emit their
motile filaments more readily there than in vitro. It was necessary now
to seek the destination of the latter in the insect's tissues; and here the
true magnitude of the task to which I had set myself became manifest.
Manson had been able to follow the migrations of the filaria embryos
with comparative ease because they are large organisms readily distinguish-
able from the fluids or tissues which surround them. But the motile fila-
ments are exceedingly delicate bodies, the movements of which are very
difficult to follow even with the highest powers of a good microscope and

* It was easy to distinguish those which had produced the filaments by their collapsed
condition afterwards.

i6

in the clear spaces of an ordinary preparation of blood. But the blood
in a mosquito's stomach shortly becomes a thick grumous mass in which
it is impossible even to see the filaments unless they are in active move-
ment. Moreover, even this assistance was denied me; for I speedily ascer-
tained that within a few minutes of their escape they seemed to lose
their movements. At least they constantly disappeared as if by magic;
and in spite of all artifices I failed to ascertain what had become of them.*
In fact to trace them further, to follow the migrations of these well-nigh
invisible bodies through the masses of cells of which so large an animal
as a mosquito is composed was indeed an impossible task with the means
which I possessed.

Hence, though Manson then and later constantly advised me in his
letters to adhere to the plan of following the motile filaments, I determined
to abandon the quest and to employ other methods; and he himself failed
to obtain any results with the insects which I sent to him from time to
time. It was most fortunate that I came to this decision so early, because
events have proved that the motile filaments migrate nowhere, and do not
enter the mosquito's tissues at all.

The first method which I now adopted and which ultimately led to
success was the following. By hypothesis, the motile filaments after reach-
ing the particular cells of the mosquito for which we thought they were
destined, should grow in them into some unknown but larger form. It
was impossible to predict what this form would be. Manson conjectured
that it would very likely be some intracellular form similar to the intra-
corpuscular stages of the organism in the human blood — fixed perhaps
in the stomach cells or blood cells of the insect. Personally however,
while I thought this view possible, I had no full faith in it. It seemed
to me that the mosquito stage of the parasite might be anything, so long
as it was of a protozoal character.

The protean changes of many of the parasitic worms warned us that
nature was capable of ordering any extraordinary transformations in the
interest of parasites; and as no case was yet known of a protozoon ca-
pable of wandering from one species of host to another, I had no guide
as to what might happen with the organisms which I was studying, and
conjectured that for all we could say, the motile filaments might develop

* It might now be possible, though still difficult, to follow them by staining them
either in dehsemoglobinised blood or in section ; but the Romanowsky reaction was not known
lo me at that time.

17

into almost any form — amoeboid, coccidiform, gregarinoid, or even in-
fusorial, small or large. What was nearly certain, however, was that they
were likely to grow in size after a few days' residence in the mosquito —
to become more visible, and, if they were to pass into the water as we
thought they would do, to take a definite form of resistance which ought
to be easily recognisable. My new method then was to give up the attempt
to follow the newly escaped and almost invisible motile filaments, and to
dissect the mosquitoes, not at once, but after the lapse of some days,
during which time the motile filaments should by hypothesis develop into
something more tangible. I proposed then to feed the mosquitoes as be-
fore on cases with crescents in the blood; to keep them alive for some
days; and then search their tissues for any parasites which might occur
in them. The parasites found, it would be easy to determine whether or
not they are derived from the motile filaments, simply by ascertaining
whether or not they also occur in mosquitoes of the same kind fed on
healthy blood. Throughout the investigation it was of course necessary
to employ only what I called in bacteriological parlance »sterile mosqui-
toes>), that is mosquitoes freshly hatched from the larvae in captivity and
therefore not contaminated by previous feedings.

Such was the procedure now adopted; but the difficulties involved
■even in it were very great. As the situation of the sought-for parasites
could not be indicated with any certainty, it became necessary to search
for them through all the tissues of each insect examined — to scrutinise
hy a powerful microscope, one by one, all the minute cells composing the
huge aggregate of which the insect consists.* To investigate a single in-
sect thoroughly in this manner required at least two hours' exhausting
and blinding work. Added to this difficulty, I had no clue as to the form
.and appearance of the object which I was seeking for; nor was I even
sure that the kind of insect under examination was amenable to the in-
fection at all. I was looking for a thing of which I did not know the
appearance in a medium which I did not know contained it. In short it
was a mere blind groping for some clue which I trusted fortune would
give in the end. As an instance of the difficulty of such work I may mention
that neither the organisms of yellow fever, which is now known to exist
in a particular kind of mosquito, nor the Pyrosoma of Texas cattle-fever,
which is known to exist in a tick, have yet been found in these animals.

* Under a magnification of a thousand diameters a mosquito appears as large as a
horse.

Researches oft Malaria. 2

though long searched for by competent observers. Nevertheless, I am
confident that, hopeless as the method may appear, it was the only one
capable of solving this difficult problem.

At the outset of the investigation it was necessary for me to become
thoroughly acquainted with the normal histology of the mosquito — for
which I had again to trust to my own observation; in spite of all efforts
no literature on the subject could be obtained by me. It was also neces-
sary to note and study the ordinary parasites of these insects — of which
I found a number during the ensuing years. Indeed, at the commence-^
ment of the work I found one which required careful working out. It
was a pseudo-navicella occurring in the malpighian tubes of the brindled-
mosquitoes iStegomyia). After a little study it was ascertained that pseudo-
navicellse have no connection with the parasites of malaria, being the
sporocysts of a species of gregarine. Next year Manson published an.
account of these interesting organisms taken from my letters to him [26].
I refer to them also in my publication at the end of the year [24].

The second method alluded to above was based on the following
considerations. According to Manson's more advanced hypothesis, the
motile filaments, after living some days in the mosquito, would probably
pass from it into the water, on the surface of which we then supposed it
usually died after laying its eggs. Such water then ought to be infective
to human beings, either when ingested, or perhaps when inhaled as a
vapour. It would be easy to test this speculation by experiment. I caused
a number of mosquitoes, both of the brindled and grey varieties, to feed
on a selected patient, and then kept them in large jars containing water
at the bottom, until they died one by one. The water was then exposed
to sunlight and otherwise allowed to remain in the condition of marsh
water. Different batches of fed mosquitoes were introduced into the jars
from time to time so as to make sure that the water should indeed con-
tain the parasites which by hypothesis should escape from the insects.
In May 1895 I gave draughts of this water to three natives who volunteered
themselves for the experiment. All of them declared that they had not
suffered from fever for years.* Strangely enough one of the men developed
a mild but marked attack of fever in eleven days, the parasite being found
in his blood. I was naturally much pleased with the success of the ex-
periment and began to hope that the mode of infection had been found;.

* The experiment was justifiable owing to the slight degree of illness usually produced
by malarial fever in natives when properly treated.

19

but the failure of many subsequent attempts of the same kind forced me
later to reject any definite conclusion on the point (section ii).*

II. Bangalore; i8g^ — p/. Progress of work. — Possessing abund-
ance of material together with plenty of leisure, I was now progressing
excellently with my researches (though without definite results) when I
received my first interruption. On the 9th September 1895, I was placed
on special duty by the Government of India to combat a serious outbreak
of cholera in the large town of Bangalore, and also to report on the ge-
neral condition of sanitation there. This duty was of great interest and
value to me because it afforded me an unrivalled opportunity for examin-
ing in the closest detail the general sanitation of a tropical city — an
experience which has stood me in good stead during later years.. For
four months, however, I was so busy with my new labours that I had
little time for research. Bangalore I already knew well, having indeed
made some of my first studies of malaria there when stafT-surgeon of the
town from 1890 — 93. I now easily ascertained by the light of Laveran's
great discovery that the cases of fever which I had attributed to intestinal
auto-intoxication were nothing but examples of asstivo-autumnal infection
among partially immunised natives. I found also that, as at Secunderabad,
my brindled and grey mosquitoes abounded all over the place. I dissected
a few mosquitoes as time allowed; and when my more arduous sanitary
duties began to diminish in March 1896, I found that I could give an hour
or two a day to the work. My results, however, remained constantly nega-
tive, in spite of the closest scrutiny of many mosquitoes. At the same
time I continued my attempts to produce infection by water.

In March 1896 Manson delivered the three Goulstonian Lectures at
the Royal College of Physicians in London, and again put the case of
his hypothesis in an admirable manner, supporting his arguments largely
upon my observations of the previous year [26]. He wrote to me fre-
quently for fed mosquitoes, which I sent to him whenever I could. He
also urged me to keep on the track of the fiagellated spores; advised me
to try infection experiments with the insufifiation of dried and powdered
mosquitoes, and with the vapour of an ^artificial marsh» in which fed
mosquitoes had died. These devices did not appear as promising to me

* Owing to the interest of Surgeon Major Owen, the Maharajah of Patiala had at this
time offered the government of the Punjab to employ me at his own expense to study malaria
in his dominions. The government of the Punjab, however, refused the offer.

20

as they evidently did to him. It was scarcely likely that dead mosquitoes
could do much in regard to the dissemination of malaria in nature, at
least in the form of dust, owing principally to the fact that dead insects
seldom escape the ants in the tropics. All dust, moreover, is generally
subject to the intense heat of the sun which, except in the presence of
water, must be very inimical to most unprotected organisms. The small
amount of time at my disposal was therefore devoted to the methods al-
ready attempted.

Towards the middle of the year I had made nineteen experiments
with a view^ to carrying infection by drinking water; and together with
three more cases, I described these in a publication at the end of the
year [30]. Water, in which mosquitoes fed on cases of malaria had died,
or which contained large numbers of the pseudo-navicellas of the gregarines
of mosquitoes, was given to various persons by the mouth. The majority
of the attempts were entirely negative: but nevertheless a slight but notice-
able reaction did occur in three of the whole number of twenty-two cases.
This still remains a very curious circumstance; but the facts were pub-
lished exactly as they were found, without the influence of the »personal
equations. At the end of the paper I summarised my results and decided
that the positive reactions, though interesting, were too few and too slight
to warrant any definite conclusion.* I am now inclined to think that they
may have been due to the following circumstances. The persons on whom
the experiments were made were generally low-caste Indians who required
a fee before drinking the water and also an assurance that they would re-
ceive more if taken ill. Now it is well recognised that many natives are
constantly infected with malaria and get relapses on any extraordinary
demand being made upon their systems, as by fatigue, chill, or dissipation.
I have even heard it stated by medical men possessing large experience
of natives that they can often produce fever in themselves by exposure
when they wish to do so. In this case at all events it was possible that
some of the subjects spent their preliminary fees in dissipation, thus pro-
ducing the supposed reaction after the experiments.

These results not being as decisive as I had expected from the first
experiment of the kind made in the previous year, I began to consider

* My actual words were, »While we cannot dream of stating definitely on the strength
of these experiments that there is something connected with the mosquito which is capable
of imparting fever, the three positive results are still curious and tend to be in favour of the
. truth of Hanson's theory.* Yet one of my Italian critics has attempted to prove, by ignor-
ing this passage, that I pretended to have established infection by drinking water.

21

whether some other route of infection was not possible or probable; and
it soon grew upon me that Manson's induction was exigent only as regards
the entry of the parasites into mosquitoes, and that his secondary hypo-
thesis regarding their escape from the insects and their infection of man
through drinking water was not so strong. I quickly thought of several
other routes for infection — which will be examined presently; and first I
considered it possible that the insects, previously infected from diseased
persons or possibly from other mosquitoes, might then inoculate the para-
sites into healthy persons during puncture, or might deposit them on the
skin during haustellation. It was easy to test this view immediately by
experiment; and early in August I made a small series of observations
which were published in the same paper [30].

A number of mosquitoes all bred from larvae in captivity, and of all
the kinds which I could collect (many specimens of brindled and grey
mosquitoes) were fed upon several patients with numerous parasites in
their blood. One of these patients had all three kinds of parasites in him;
and I specially employed this case, as well as many varieties of mosqui-
toes, in order to increase the chances of one at least of the species of
mosquitoes present being appropriate for one at least of the species of
parasites. After feeding, the insects were kept alive for one or two days
and were then applied in considerable numbers on two occasions to Mr.
Appia, Assistant Surgeon of the Bowring Civil Hospital at Bangalore, who
courageously volunteered for the experiment. Mr. Appia had suffered
from malarial fever some years previously, but not .since then; so that if
he should be attacked by fever shortly after the experiment, it would be
strong evidence, if not proof, in favour of the inoculation theory. He re-
mained, however, absolutely free from fever. He was then bitten by five
mosquitoes which had been partially fed immediately before on a case of
crescents — on the supposition that the insects may carry the infection
mechanically, as the tsetse fly carries nagana; but the result was again
negative. Lastly two other individuals were bitten by mosquitoes fed from
three to five days previously; still without effect. I judged then, either
that infection is not produced in this way, or that the proper species of
mosquitoes had not been employed, or that they had not been kept for
the proper period after feeding; and I proposed to return to the subject
again. It should be noted that these experiments of mine were made
quite independently, and before I had heard of the theories of King and

22

Bignami — as indeed was stated in another publication of mine at the end
the year [32, p. 251].

In July 1896 Bignami's criticism of Manson's hypothesis, referred to
in section 6, appeared in Italy [29]. I heard nothing about it whatever,
until I received Manson's letter of the 12th October, which was accom-
panied by a translation of the critique. Bignami's paper was not a pro-
found one, and consisted only of a copious and dexterous rendering of
ideas which were new only to those who had not already fully considered
the subject. His objection to Manson's theory was based principally on
Grassi's loose speculation that the motile filaments are the result of the
death of the parasites in vitro. As this was a vital point in the chain of
reasoning I now set to work to examine the subject experimentally, and
was soon able to show that the escape of the filaments depends on certain
proper conditions, and not at all on the death of the parasites. Thus
they escape more readily when the specific gravity of the blood is al-
tered, either by the abstraction of water by partial evaporation or, as
Marshall proved, by the addition of a little water. On the other hand
they do not escape at all, even when the parent cells perish, so long as
the blood is kept scrupulously unchanged. In order to prove this, I drew
the blood from the finger into a small mass of vaseline placed upon the
skin, and then mounted the whole for the microscope in such a manner
as to prevent the blood coming even into momentary contact with the
air. The result was that not a single crescent emitted motile filaments
or even underwent the preliminary change of spherulisation, although it
was evident they all died after a time.* This experiment completely
<lisposed of the death-agony theory of the Italians. Previously to this,
however, Sacharofif had shown that contrary to Grassi's statements, the
filaments do contain chromatin; but I could not procure a copy of his
work [23]. I should add that after long observation of the filaments I
could never bring myself to believe that they are merely the result of
the spasmodic movements of dying protoplasm; and this tale was in fact
never anything but a gratuitous assumption.

These researches were published later [32, 33], and were confirmed by
Manson and Rees in London [37]-

* If the preparation was opened and the blood momentarily exposed to the air within
some hours after abstraction from the patient, the crescents could be seen at once to resume
their functions. But if this experiment was delayed about 24 hours, the crescents no longer
reacted, and indeed showed clear evidence of death by their vacuolisation and other struc-
tural changes.

23

The conditions required by the crescents for emitting filaments were
now clearly seen to be those obtaining in the mosquito's stomach, where
the blood is rapidly altered by abstraction of water; and I therefore con-
tinued my work without further reference to Bignami's objection.

His view that infection may be caused by inoculation had already
been considered and experimented on by me, as just mentioned. But it
should be understood that Bignami's hypothesis (which was the same as
that given long previously and much more strongly by King) was very
■different from mine. King and Bignami thought that mosquitoes bring
the poison from marshes to man; this speculation had not occurred to me
until I read Bignami's paper in October, and then it did not appeal to
me at all, because it was self-evident that the connection between malaria
and marshes could be sufficiently explained by the fact that mosquitoes
breed in stagnant water. My speculation was that mosquitoes become in-
fected from men (according to Manson's induction) and possibly also from
■other mosquitoes, and then communicate the parasites to healthy persons
— perhaps by inoculation. It will be seen which view is right; but in
■consequence of my negative experiments, the inoculation theory was not
much favoured by me until I made my researches in the Sigur Ghat
{section 12).

My duties at Bangalore continued for a year and a half. At first
placed upon special duty to report on the sanitation of the town (80,000
inhabitants), I was afterwards appointed officiating Residency Surgeon there
and was required to reorganise the whole of the sanitary arrangements,
to create a health department, to participate in a committee designated
to reconstruct the municipal regulations, and to contend against several
outbreaks of cholera. Consequently I did not possess as much time as at
Secunderabad for my researches on malaria, but nevertheless in addition
to the experiments last referred to, I was able to dissect many hundreds
of mosquitoes in pursuance of my principal plan of campaign. Several
agents were employed to collect the larvae of as many kinds of mosquitoes
as possible, especially from several spots whence most of the cases of
fever came; and these insects, belonging to many species of the brindled
and grey groups of mosquitoes,* were all tested by direct feeding on cases
of malaria, especially sestivo-autumnal. But though each insect was exa-

* Not once as yet had I come across the dappled-winged mosquitoes; though, be it
noted, many of my larvse were collected from ditches and the edges of ponds.

24

mined with the utmost care, almost every cell being scrupulously searched
for parasites, the results still remained entirely negative.

Towards the end of my stay in Bangalore, as failure followed failure,.
I was naturally forced to reconsider the whole basis of my work. But no;
the most critical examination of Manson's induction failed to exhibit an}'
flaw in the fundamental reasoning. The gametocytes, and the process by
which the motile filaments escape from them after the blood is drawn
from the patient, could only be meant for infection of the mosquito. There
was no other explanation. Nature does not create these complex phenomena
for nothing; and the theory must be — was — sound. What then was
the cause of my repeated failures? Was it possible that the kinds of
mosquitoes which I had tested hitherto — very many kinds — were all
of the wrong species?

The reasons for and against this view were as follows. In all the
districts and towns of India in which I had served or stayed during fifteen
years — Madras, Bangalore, Moulmein, the Andamans, Secunderabad.
Upper Burma, Bombay, Poona, Calcutta, Karachi, Quetta, the Nilgherr}'
Hills, malaria was undoubtedly present, especially among the natives; and
in all of them without exception I remember to have noticed mosquitoes
belonging to both the grey and the brindled classes. This naturally sug-
gested a connection between the disease and the insects; but, on the other
hand, were not the latter perhaps too common? So far as I could ascer-
tain, the disease Avas generally limited to certain spots and localities (by
no means always near marshes); whereas the insects were everywhere,
and were indeed often commonest at points were malaria was rare, as in
the houses of Europeans. After all may not the true malaria-bearing variet}'
or varieties have been overlooked by me? Possibly they were comparatively^
rare species, or species occurring only at a certain season — a hypothesis-
favoured by the well-known fact of the seasonal variation of malaria.
Now, as I was fully aware at the time, malarial fever is a relapsing dis-
ease in which attacks continue to occur for years after infection; so that
it does not follow by any means that the infective variety of mosquito
must always be present in a locality, even though numerous cases of malarial
fever are present. And it was to be specially noted that most of the cases
occurring in Bangalore were probably only cases of relapse.

These arguments were not strong enough to be conclusive on either
side of the question. I had done quite right in spending so much time
over the grey and brindled mosquitoes; there was enough prima facie

25

evidence against them to demand a full enquiry. But before spending-
more time over them it was now advisable to see whether further light
could be obtained by epidemiological investigation. The towns in which
I had worked hitherto could scarcely be considered more than moderately
malarious; I now proposed to visit an intensely malarious spot, at the
height, too, of the malarious season in order to ascertain what kind of
mosquitoes prevailed there at the time; and reasonably hoped that this kind
would prove to be the guilty species.

Being a servant of Government I could not of course go where I
pleased without leave, and I therefore first attempted to interest Govern-
ment in my work. Owing to my representations, the United Planters'
Association of Southern India took up the matter; and the Honourable Mr.
Bliss, Member of Council of the Madras Government, and also Surgeon General
Sibthorpe, head of the Madras Medical Service, were kind enough to give
their warm assistance — for which I shall always be much indebted. The
result was that the Government of Madras made a proposal to the Govern-
ment of India that I should now be placed on special duty to investigate
malaria. Most unfortunately, however, in addition to the plague, the Afridi
war broke out just about that time, and owing to the paucity of medical
officers the Government of India was obliged to reject the proposal —
May 1896. But in the meantime I had determined to begin the enquiry
at once at my own expense during two months' leave which was due to
me; and accordingly, on the completion of my duty in Bangalore, I went
to the Nilgherry Hills for the purpose of studying the point referred to
in some of the intensely malarious plantations at the foot of these mountains.

12. 77/1? Sigur Ghat; iSgy. — I arrived at Ootacamund, the great
hill station of the Nilgherry Hills, at the beginning of April, 1896. This
station which is about 8,0(X) feet above sea-level, is surrounded by nu-
merous tea and coffee plantations, scattered here and there in the rich
valleys of the hills, and even for some distance out on the plains which
encompass the hills like a sea. A.fter enquiry it was determined to begin
the investigation in the Sigur Ghat, a long natural trench which cuts at
one stroke from the Ootacamund plateau right down to the plain, and
which had the worst reputation for malaria. A dak bungalow (rest house)
and a small plantation existed near the top of the trench, at a place call-
ed Kahutti about 5,500 feet above sea-level; and owing to the fact that
a single night spent lower down the valley was thought enough to ensure

26

a bad and perhaps fatal attack, I determined to lodge here and visit the
lower valley only during the day time. Nevertheless even at Kalhutti I
found almost everyone suffering from fever — which was ascribed to
miasmata floating up the ravine from the plains below; and I had been
there only a few days and had paid only one diurnal visit to the plain
when I myself suffered a bad attack of aistivo-autumnal infection, the
diagnosis being confirmed by the microscope.*

After two weeks' energetic treatment with quinine I was well enough
to resume operations; and this time went direct to the plantations at the
foot of the Sigur Ghat. The owner of one of them, Mr. Kindersley, wise
enough to reside in the hills during the intensely malarious season of the
year, very kindly placed his house in the plantation at my disposal; so
that I was able to make a thorough survey of the locality. Both planta-
tions are situated in the midst of luxuriant forest and undergrowth close
under the declivities of the mountains, and are copiously watered by irri-
gation channels. Almost all the native employes^ as well as some families
of aborigines, were suffering from severe malaria — anaemia, emaciation,
and enlarged spleen; and the parasites were easily found in the blood of
some of them. But I was not a little astonished when I discovered that
mosquitoes appeared to be almost absent in all the houses. In spite of
considerable rewards which were offered for their capture, and in spite of
the efforts of my trained servants and myself, scarcely any were secured.
T was informed indeed by some of the employes that they were often bitten at
night by insects which escaped in the morning; but these nocturnal visitors
were not procurable.** Later however, we were told of some insects
which haunted the jungle and bit in the daytime under the trees. I found
these to be a small kind of brindled mosquito, and strongly suspected
that they might be the culpable species; and accordingly examined them
closely and called them Cttlex silvestris.

A part of my mission here was to enquire whether the mosquitoes
in this highly malarious spot did not contain parasites which w'ere not con-

* This case was remarkable for the brevity of its incubation period. I had never suf-
fered before from malaria, and was not likely to have acquired the infection either at Bangalore
or Ootacamund. I had arrived at Kalhutti at 6 p. m. on the 22nd April, and my attack
commenced at lO p. m. on the 25th April. I ascribed it at the time to my visit to the plain
made on the 23rd April; but there is now little doubt that the infection was acquired at
Kalhutti itself, which was swarming with mosquitoes, and where the servant of the dak bunga-
low and all his family were ill. At the same time I do not remebember to have been bitten
by mosquitoes, and said so in my published account.

** Judging by our present knowledge, these must have been the offenders.

27

tained in the mosquitoes of the less malarious Bangalore. If they did so
these parasites might reasonably be suspected of being the mosquito stage
of the malaria parasite; and the question could subsequently be tested by
■experiment. These mosquitoes were at once found to contain two new
Tiinds of parasites, namely crowds of active swarm-spores in the intestine,
and, secondly, clusters of spores (each cluster containing eight bright oval
spores) in the ventral nervous system. A close study was made of these
■organisms; but they did not appear in some of the jungle mosquitoes
which had been fed on patients. Strangely enough however, a person who
volunteered to swallow a number of the swarm-spores in water was attacked
subsequently with fever, the malaria parasites, however, not being found
in his blood; but I heard afterwards that, contrary to his statements, he
had had fever just previously.

It will be remembered that Manson's secondary hypothesis suggested
that the motile filaments, after living for some time in the mosquito, pass
from it into the water, and thence by ingestion or inhalation into man. M}'
experience, however, tended to convince me that if such infection of water
takes place at all it must be very limited — in other words, that after
their escape from the dead mosquito, the organisms can neither travel
far in the water nor live long there. For if they could do this, almost
all water in India would be infected, and the disease would be universal,
instead of being confined, as it is, to certain spots. For the same rea-
son the miasmatic theory never appealed strongl}- to me. I thought it
most likely that men became infected from small stores of drinking water
such as wells, cisterns, and even pots and ewers, into which infected mos-
quitoes often fall and die while laying their eggs — a theory which would
•easily account for the isolation of the malady, because, as I had observed
over and over again, mosquitoes seldom wander far from their haunts.
As, according to hypothesis, the organism escape from the gnat into the
water in which she lays her eggs, it followed that water which contained
most larvae should contain most malaria parasites, and, conversely, that
■drinking water free from larvae would probably be free from parasites.
Now in attempting to apply these considerations to the case of the Sigur
plantations, I found them at once opposed by many facts. Not only were
there few adult mosquitoes there, but the larvse could be found only in
a few stagnant puddles in the depth of the jungle, while the drinking
water was obtained from rapid streams just issued from pure mountain
springs, in which larvae neither existed nor were likely to exist.

28

These facts again forced me to reconsider the whole of Manson's se-
condary hypotheses, and to search for more plausible theories. Three such
theories occurred to me. I had long observed that while they are sucking
blood, gnats deposit minute drops of excretae on the skin every ten se-
conds or so; and I had actually shown that these drops may contain the
pseudo-navicellas of gregarines. It was therefore possible that they might
contain the spores of the parasites of malaria, which might then be able
to work their way through the skin and into the blood of the victim.
Another hypothesis of mine was that the malarial spores might be voided
by the insects, not upon the skin, but upon rotting vegetation or damp
earth (e. g. the floor of the houses and huts of natives), and might there
possibly develop into some extracorporeal form capable of infecting man
by air-borne spores.* The third theory was that infected mosquitoes could
in some mysterious manner introduce the parasites directly into the blood
during the acts of puncture and haustellation. This view was similar to
that of King and Bignami, with this difference that while these observers
thought that the mosquitoes derived the parasites from marshes, I held, in
consequence of Manson's induction, that they derived them from patients.
In the account of my work in the Sigur Ghat which was published a few
months later [40] it was stated that this was the hypothesis Avhich I now
held to be the probable one.**

It was during these researches that I first noticed the »dappled-winged
mosquitoes». While looking for mosquitoes in a vacant rest house at the

* This was by no means an idle conjecture, and was indeed strictly based upon the
analogy of Cunningham's life history of the Amoeba eali, which that observer stated was voided
from the intestines of cattle and afterwards formed pseudo-plasmodia in the exposed dung — men
and cattle being infected by the air-borne spores of these pseudo-plasmodia. He thought that
the organisms were related to the Mycetozoa and called them Proiomyxomyces coprinarius.
His important statements have been ignored but not disproved by subsequent writers. Simi-
larly I thought that the parasites of malaria might possibly be extracted from the circulation
by mosquitoes, be deposited by them upon the damp floors of dwelling houses and there de-
velop in a like manner. This hypothesis was at that time as cogent as any other.

** I said, «0n the whole from a consideration of the epidemiological facts I should be
inclined to favour the idea of contact being the mode of infection ; and may add that one of
my servants who was employed in catching the adult silvestris by allowing them to settle on
his legs and arms was attacked five days afterwards by the quartan parasite. « By contact I
meant contact of the mosquito with the skin as explained further on by the following words :
'Since the presence of a human being in the jungle at once causes a number of silvestris
mosquitoes to attack him on all sides, it is very clear that he may readily be infected by
their agency, either by injection of the parasite through the puncture, or by its deposition on
the skin in the shape of spores contained in the insect's faeces, which, observation shows, are
always discharged in quantity during the act of haustellation. « My theories regarding infec-
tion are also referred to in my previous paper [30].

29

foot of the ghat I captured an insect resting in a peculiar attitude with
the body-axis at an angle to the wall (as I noticed at the moment). On
examination, its wings were found to have a series of black marks along
the anterior nervure; but as I saw no more individuals of the species, I
did not think the observation to be of sufficient importance to be included
in my paper. Yet, had I only known it at the time, this was the very
•species I was in search of!

Indeed the whole of this investigation afforded a clear example of
the well-known ambiguity of epidemiological work. Of the kind of insect
which was really causing the disease at the time, I saw but a single indi-
vidual! The reason is now quite apparent. Unlike the grey and brindled
mosquitoes which rest in the dark corners of dwellings by day in large
numbers, many species of dappled-winged mosquitoes fly out at daybreak.
It is true that other species of this genus have more domestic habits and
can therefore be more easily found; and if fortune had been my friend
in those days she would have brought me to a place where these species
abound — such as places afterwards visited by me in Assam and the Dar-
jeeling Terai. Nor does it follow in any case that the predominant spe-
cies of mosquito in a locality must be the malaria-bearing species there;
there is no reason why the innocent species should not outnumber the
■dangerous species even in the most malarious spots: while lastly, it is
now known that the dangerous species may abound where there is no
malaria at all. Hence, though I did not know it at the time, it is im-
possible to indicate, much less to certify, the malaria-bearing species by
its numerical relations with other species in malarious localities.

One of the principal results of my work in the Sigur Ghat was that
it led me to doubt the probability of infection by drinking water. I should
have liked to remain there much longer; but on the expiry of my leave
was forced to return to my regiment at Secunderabad, five hundred miles
away, and was never able to visit the place again.*

13. Secunderabad; iSgj. The Fundamental Discovery. — On my
return to Secunderabad (July 1897), ^he first thing I noticed was that the
malaria had continued unabated during almost two years since I had left; if
anything it was worse, and many recruits who had recently joined the regi-
ment had been attacked — as they averred, for the first time. This clearly

* I had been offered an appointment in Berar, but had declined it in order to carry
on these researches in the Sigur Ghat. I suffered severely for this later on.

30

showed that these cases were not merely relapses, and that some cause of in-
fection was actually at work among the troops. It was for me to discover the
cause; and I determined to return to my old method, and to test experimentally
all the kinds of mosquitoes prevalent anywhere near the barracks. I had
now been studying the subject almost constantly for over two years, and
had become so very familiar with the microscopical appearance of the
various structures of the mosquito* that I felt the mosquito stage of the
parasite could no longer escape me if it existed at all. Numerous »cases
of crescents* suitable for the experiments were in my hospital, and it was
obvious from the number of fresh cases occurring that the proper kind
of mosquito must be somewhere about. If I failed it could only be be-
cause there was some flaw in Manson's induction.

At the same time a possible fallacy was detected in the logic of that
part of the theory which suggested that the motile filaments after their
escape from the parent cells in the mosquito's stomach must take up"
their abode in the tiss?/es of the insect. The vital and inevitable part of
the induction consisted only of the reasoning which inferred that the sto-
mach of the mosquito is the natural loctcs for the escape of the motile
filaments. It was only conjecture to say that they must enter the tissues;
because for all we knew it was possible that they might remain in the
intestine for some time and then be voided, probably in some altered
form, either upon the ground or upon the human skin (see my hypo-
theses in the previous section). It was therefore now necessary to examine
the evacuations as well as the tissues of my subjects.

I commenced work by making a careful survey of the various kinds
of mosquitoes which were to be found in the officers' quarters, in the
regimental hospital, and in the numerous little houses of the native sol-
diers, which constituted the barracks or »lines», as they were called. I
found first, the insects with which I was familiar during my previous
studies here in 1895, namely (a) several species of brindled mosquitoes,
and (b) two species of grey mosquitoes. But at the same time I was
astonished at observing that the whole place was overrun by swarms of
(c) a small and delicate variety of mosquitoes, which were at once ob-
served to rest with the body-axis at an angle to the wall, and which had
spotted wings. In fact they were evidently of the same genus (though
not of the same species) as the mosquito which had been previously found

* This does not mean that I was equally familiar with the niacroscopical anatomy of
the mosquito — a subject which has only recently been dealt with fully.

31

in the Sigur Ghat — a genus, or perhaps family, quite distinct from those of
the grey and brindled mosquitoes with which I had hitherto been working.

It is now time to speak more particularly of all these mosquitoes.
I had written repeatedly to Manson, to various booksellers in England,
and to several persons in India who I thought might help me, for some
literature on the subject; but could obtain nothing except a few notes by
popular authors, such as Thomas, who wrote on piscatorial subjects in
India. I could not even obtain any adequate works on the anatomy of
insects in general. Of Ficalbi's work on European gnats — which would have
helped me immensely — I was ignorant, and received no copy. Manson
had found the name of one species of mosquito which I sent to him; but
this did not help me, for what I required was a scientific work on the
structure and classification of the mosquitoes as a group. I was therefore
obliged, as mentioned in section lo, to trust to my own rough methods
of classification; and these were based, not on the criteria of entomologists,
such as the structures of the mouth parts or the nervures of the wings,
but on the general appearance and markings, the eggs, the habits, etc.,
of the insects. It was only the working classification of an amateur
without literature to guide him, and made for his own convenience; but,
as events have proved it was roughly correct. Up to July 1897 I recog-
nised the two following groups: —

(a). Brindled Mosquitoes (now recognised as belonging to the genus
Stegoniyia, Theobald). Body and legs boldly marked black and white, or
brown and white. Wings plain. Biting voraciously, mostly in the day-
time. Resting with abdomen hanging towards the surface of attachment,
and the last pair of legs tilted on the back. Breeding mostly in pots of
water. Larvae floating head doAvnwards and possessing short stumpy
breathing tubes. Eggs black, oval, and laid separately.

(b). Grey Mosquitoes (now recognised as belonging to the genus
Culex, Linn, as defined by Theobald). Back barred with transverse brown
and white stripes. Legs and wings plain. Biting somewhat timidly, mostly
at night. Resting with abdomen hanging towards surface of attachment.
Breeding mostly in wooden tubs, ditches, garden cisterns, and drains.
Larvae floating head downwards and possessing long breathing tubes.
Eggs elongated and somewhat lanceolate and laid simultaneously in »rafts.»

I had found mosquitoes of the same genera, though possibly of dif-
ferent species, at Bangalore and at several spots in the Nilgherry Hills;
and also at Bombay, Poona, and Madras during short visits made to these

32

cities in connection with my sanitary duties at Bangalore. I remembered
also to have seen similar insects in Burma and the Andamans; so that it
was reasonable to suppose that the}' constituted the common or ordinary
kinds of mosquitoes in India. The new mosquitoes which I now and
subsequently met with, and named dappled-winged mosquitoes, were evi-
dently of quite another genus to the foregoing, and were distinguished
by me by the following characteristics: —

(c). Dappled- Wiiiged or Spotted- Winged Mosquitoes (now recognised
as belonging to the genus Anopheles Meig.). Body, legs, and proboscis
marked brown and white, or dark and light brown. Wings with several
dark blotches on or near the anterior nervure. Resting with abdomen
pointed outward from the surface of attachment. Body more elegant,
and shaped like that of a humming-bird moth. Breeding mostly in na-
tural pools of water on the ground. Larvae floating flat on the surface
of the water like sticks, and possessing no breathing tube at all. Eggs laid
singly; cohering in triangular patterns, and shaped like an ancient boat with
raised prow and stern, and surrounded with a membrane which — when the
^gg is seen in profile — gives the appearance of a bank of oars to the boat.

In the spotted-winged mosquitoes which I now found at Secunderabad
I noticed at once the general difference of shape, the peculiar attitude of
the insects when at rest, the marks on the wings, and the appearance of
the eggs (as seen within the body of the female when dissected); but
the larvae could not be studied until later.* The adults were very deli-
cate, pale brown creatures, which by common consent seemed scarcely
to bite man, though they were numerous enough to have caused much
irritation had they done so. They swarmed in my own quarters, but
seldom bit me. They abounded also in the houses of the other officers
of the regiment, who, with their families had remained quite free from
malarial fever. Consequently I was not disposed to think that they had
anything to do with the disease. On the other hand the grey mosquitoes
swarmed in the barracks, but were much less numerous in the officer's
quarters (situated some hundreds of yards to leeward of the barracks).
Suspicion therefore first attached to the latter variety.

I determined, however, not to be swayed by such considerations, but
to make a most complete and exhaustive test of all the varieties which I
could procure — even at the cost of repeating much of my old negative

* It was principally my assistant, Mahomed Bux, who ascertained, as a general rule, the
attitude of the larv?e.

33

work, during which, laborious as it was, I may have overlooked the object
I was in search of. A number of natives were employed to collect larvae
from far and wide round the barracks. These larvae were kept in sepa-
rate bottles, and when the adult insects appeared they were released with-
in mosquito nets in which the patients were placed. The insects were
applied sometimes during the day in a darkened room; and were some-
times fed all night. After feeding, the gorged insects were collected in
small bottles containing a little water and were kept for several days be-
fore being dissected. The procedure was therefore the same as before;
but now, in order to ensure at least definite negative results, redoubled
care was taken; almost every cell was examined; even the integument
and legs were not neglected; the evacuations of the insects found in the
bottles, and the contents of the intestine were scrupulously searched; at
the end of the first examination staining reagents were often run through
the preparation, and it was searched again with care. The work, which
was continued from 8 a.m. to 3 or 4 p.m. with a short interval for break-
fast, was most exhausting, and so blinding that I could scarcely see after-
wards; and the difficulty was increased by the fact that my microscope
was almost worn out, the screws being rusted with sweat from my hands
and forehead, and my only remaining eye-piece being cracked, while
swarms of flies persecuted me at their pleasure as I sat with both hands
engaged at the instrument. As the year had almost been rainless (it was
the first year of plague and famine) the heat was almost intolerable, and
a punkah could not be used for fear of injuring the delicate dissections.
Fortunately my invaluable oil-immersion object-glass remained good.

Towards the middle of August I had exhaustively searched numerous
grey mosquitoes and a few brindled mosquitoes. The results were abso-
lutely negative; the insects contained nothing whatever. Then, I think
for the first time, I began to feel that the long quest had been in vain
and that a flaw existed somewhere in the induction. The disease was
there, the mosquitoes were there — how was it that I found nothing? I
may perhaps be pardoned for dwelling on my personal feelings during
that time, and the astonishing time which followed. Science too has its
drama; and the actor on that real scene cannot help being moved when
he remembers it — although it may appear trivial enough to others.

I had remembered the small dappled-winged mosquitoes, but as I
could not succeed either in finding their larvae or in inducing the adult
insects to bite patients, I could make no experiments with them. On the

Researches on Malaria. 3

34

1 5th August, however, one of my assistants brought me a bottle of larvae,,
many of which hatched out next day. Among them I found several
dappled-winged mosquitoes, evidently of the same genus as those found
about the barracks, but much larger and stronger. Dehghted with this
capture I fed them (and they proved to be very voracious) on a case
with crescents in the blood. Expecting to find more in the breeding
bottle and wishing to watch the escape of the motile filaments in this-
new variety, I dissected four of them for this purpose immediately after
feeding. This proved to be most unfortunate, as there were no more of
these insects in the bottle, and the results as regards the motile filaments
were negative. I had, however, four of the gorged dappled-winged mos-
quitoes left; but by bad luck two of the dissections were very imperfect
and I found nothing. On the 20th August I had two remaining insects
both living. Both had been fed on the i6th instant. I had much work
to do with other mosquitoes, and was not able to attend to these until
late in the afternoon when my sight had become very fatigued. The
seventh dappled-winged mosquito was then successfully dissected. Every
cell was searched, and to my intense disappointment nothing whatever
was found, until I came to the insect's stomach. Here, however, just as-
I was about to abandon the examination, I saw a very delicate circular
cell apparently lying amongst the ordinary cells of the organ, and scarce-
ly distinguishable from them. Almost instinctively I felt that here was
something new. On looking further, another and another similar object
presented itself. I now focussed the lens carefully on one of these, and
found that it contained a few minute granules of some black substance-
exactly like the pigment of the parasite of malaria. I counted altogether
twelve of these cells in the insect, but was so tired with work and had
been so often disappointed before that I did not at the moment recognise
the value of the observation. After mounting the preparation I went
home and slept for nearly an hour. On waking, my first thought was
that the problem was solved; and so it was.

Next morning I returned to the hospital with much apprehension lest
the eighth and last dappled-winged mosquito should have died and be-
come decomposed during the night. It was alive; and was killed and
dissected with much anxiety. Similar bodies were present in it, only
they were distinctly larger. The seventh mosquito had been dissected
four days after finding; the eighth five days after feeding; the parasites-
in the latter had lived a day longer than those in the former and were=

35

consequently larger. Both insects had been bred from larvae in captivity;
both had been fed for the first time on the same person — a case of ma-
laria; no such objects as these pigmented cells — as I then called them —
had ever before been seen in the hundreds of mosquitoes examined by
me; the objects lay, not in the stomach cavity of the insects, but in the
thickness of the stomach wall; all contained a number of black granules
precisely similar in appearance to those contained by the parasites of
malaria, and quite unlike anything which I had ever seen in any mos-
quito previously. Lastly, these two mosquitoes were the first of the kind
which I had ever tested.*

The mind long engaged with a single problem often acquires a kind
of prophetic insight, apparently stronger than reason, Avhich tells the
truth, though the actual arguments may look feeble enough when put
upon paper. Such an insight is mainly based, I suppose, on a concentra-
tion of small probabilities each of which may have little weight of itself;
but in this case at all events the insight was there and spoke the truth.

These two observations solved the malaria problem. They did not
complete the story, certainly; but they furnished the clue. At a stroke
they gave both of the two unknown quantities — the kind of mosquito
implicated and the position and appearance of the parasites within it.
The great difficulty was really overcome; and all the multitude of import-
ant results which have since been obtained were obtained solely by the
easy task of following this clue — a work for children. We may rest as-
sured that if these observations had not been made we should still have
remained ignorant of the mode in which this important disease, with its
annual death roll of millions, is propagated— aye, and would have remain-
ed ignorant of it until some one else had taken up the same investiga-
tion by the same method.

And no other method would have solved the problem. It was ne-
cessary to find not one but two unknown quantities, and neither could
be found by itself. There are no phenomena which would serve to indi-
cate the kind of mosquito. In nearly all malarious places there are many
kinds of mosquitoes, and, as in the Sigur Ghat and other places, the ma-
laria-bearing species are in no way predominant among them either in

* On the assumption that these cells had developed from the motile filaments it was
difficult at the moment to explain the pigment within them — as the motile filaments have
no pigment. I thought it possible, however, that after fixing themselves in the stomach wall
they might be able to derive haemoglobin from the contents of the organ, and afterwards
convert this intjD the pigment.

36

numbers or in any other way. Indeed the malaria-bearing species occur
in places where malaria has not been known in the memory of man, as
around Liverpool. By what process of reasoning then could we isolate
the species? It might possibly have been practicable to detect it by a
very long series of experiments aimed at infecting men by the bites of
successive species of mosquitoes; but no one would have undertaken such
a work without the guide of a very strong theory in favour of inoculation
by the bite; and the theory of King and Bignami to this effect was little
more than a conjecture. It was not likely that the first species tried
would have given successful results, as my own experiments of 1896
showed. Even if, after a multitude of costly and dangerous experiments, a
positive result had been attained by this method, it would always be open
to doubt (seeing that the experiments would have to be done in a mala-
rious country) whether the case was not merely one of relapse; and an-
other long series of experiments would be required to eliminate this doubt.
And then, even when the proper species of mosquito was detected, there
would still be no guide to the form and position of the parasites within
it, or even to the way in which they enter the insect (Bignami thought
that they enter the larvae from marsh water). No, the thing was not
practical. Bignami himself abandoned his experiments on his own theory
after the first failure [29] and did not resume them until after my work had
clearly indicated both the kind of mosquito implicated and the route of infec-
tion. The only practicable method was to attempt to find both unknown
quantities simultaneously by the »trial and failure system» — such as I adopted.*

The discovery of the pigmented cells, therefore, ended for me at
least the old research, the period of doubt, the groping in the dark. The
secret spring had been touched, the door flew open, the path led onward
full in the light, and it was obvious that science and humanity had found
a new dominion. But it was necessary to follow the clue forthwith; to
watch the development of the pigmented cells in mosquito after mosquito;
to ascertain what became of them; to fathom the mystery of the route
of infection; and then — to save human life in the gross, perhaps to open
continents to civilization.

The first thing was to obtain more — hundreds — of these large dap-
pled-winged mosquitoes. Alas, the man who had found them had, contrar}'

* I mention these facts because many writers on the subject seem to think that the
original discovery was made merely by catching the first mosquito and finding the pigmented
cells within it.

37

to my orders, put the larv^ from many sources in the same bottle! AH
the larvee from all these sources were collected — but no more dappled-
winged mosquitoes! I turned then to the small but similar variety which
swarmed about the barracks. Being evidently of the same genus, they
too would probably harbour the parasites; but though my men and my-
self searched high and low for their larvae, we could not find them. I
could scarcely even persuade the adults to lay their eggs in captivity.

Thinking that in spite of all my care I may have overlooked the
pigmented cells in the grey and brindled mosquitoes, I now searched for
them in the stomachs of a number of these, but without result. A num-
ber of the small dappled-winged mosquitoes caught about the hospital
were also examined for them in vain. These observations served however
for a ;>control» on the two positive cases.

Owing to the great heat at Se-
cunderabad I had been obliged to leave
my family at Ootacamund, and was
now compelled to go to Bangalore
for a few days in order to settle
them there for the remainder of the
summer. This gave me leisure for
writing a report to the government
of India on the discovery of the pig-
mented cells, and also a short paper
on the same subject for publication.

The latter was of course intended quitoes {Anopheles). From Ross's paper, British

1 1- • , 1 . -1 1 Medical Journal, i8th December 1897,

only as a prelimniary to a detailed o

■^ r J p^gg 17S7.

report which I hoped to be able to pu-
blish in a few months and which I thought would contain the full explica-
tion of the whole problem. I described my method in a few opening
lines, being careful to note that the mosquitoes used by me had been
»bred in bottles from the larva:>. The mosquitoes were then described
as well as possible — the spots on the wings and the peculiar shape of
the eggs being noted, but reference to the peculiar attitude being inad-
vertently omitted. Next I gave in detail the circumstances under which
the pigmented cells were found, together with a description of them; and
finally discussed, very guardedly, their probable relation to the parasite
of malaria. I had brought the original preparations with me, and now
showed them to my friend Surgeon Major John Smyth, who at my request

••V

Figure 2. Pigmented Cells (zygotes) of aes-
tivo-autumnal parasite in dappled-winged mos-

38

kindly added a note to my paper, corroborating my description. They
were then despatched by post to Manson. My paper, however, did not
appear until December [38]; but when it did so it was accompanied by
an excellent drawing of the pigmented cells furnished at the instance of
Manson, and also by remarks of Manson, Bland Sutton, and Thin, who
discussed the new objects — the last holding that the cells were ordinary
cells of the stomach wall into which malarial pigment had entered in
some manner from the stomach cavity. This preliminary article was
published by me for the express purpose of guiding the researches ot
others; and in fact anyone who had read my description of the pigmented
cells and of the dappled-winged mosquitoes would now have had little
difficulty in repeating my work.*

On my return to Secunderabad I was much disappointed to find that
the larvae of neither the large nor the small species of dappled-winged
mosquito had yet been collected. Consequently in the intervals of search-
ing for them, I spent my time in examining the stomachs of all the mos-
quitoes I could catch for the pigmented cells. I hoped especially to find
them in the small dappled- winged insects caught about the hospital, where
there were several cases of malaria, but was disappointed. On the i8th
September, however, a large grey mosquito was observed feeding on a
patient suffering from the benign tertian parasites and was promptly se-
cured. The stomach was full of black blood, so that it must have fed
previously (freshly imbibed blood showing red in the insects) as well as
on this occasion. It was kept until the 21st and was then dissected. To
my delight the pigmented cells were again found, in considerable num-
bers; but they were larger even than those of the mosquito of the 21st
August. As this particular insect had not been bred from the larva in
captivity I could not say for certain where it had become infected, but I
thought it likely that it had been feeding on the case of tertian all the
time (that is from about a week before it was killed) as the patient was
in a bed by himself in a corner of a large nearly empty ward. Hence I
naturally inferred as a probability that the pigmented cells in this insect
were derived from that case; and I thought that their large size suggested
that they must have been so derived about a week before the insect was
killed. But of course I could not speak with absolute assurance on these
points.**

* This is exactly what was done by the Italian observers fifteen months later (see section 23).
** This mosquito also contained a number of the swarm-spores which I had observed in
the Sigur Ghat.

39

Meanwhile swarms of small grey larvse had been found in an isolated
^ool of rain-water, which I had overlooked because it was on the top of
a hillock where pools were not likely to exist. On hatching out, these
were found to be the long-sought larvae of the small dappled-winged
mosquitoes. I observed at once that they had no breathing tubes and
that their attitude was peculiar as compared with the larvae of other
mosquitoes; and noticed also that the pool in which they were found
seemed too shallow and evanescent for the latter — facts shown by me
and my colleagues in 1899 to be of the greatest importance in connection
with the prevention of malaria. Directly enough of the adults appeared
from the larvae in the breeding-bottle, they were released in large num-
ibers within the mosquito-net of a patient with crescents in his blood.
Next morning only two of them were found to have fed themselves. One
was killed next day, but nothing was found in it. The second was killed
the day after, and was found to contain a large number of very small
-pigmented cells! This really almost clinched the matter; for three out
of four dappled-winged mosquitoes bred from the larvae in captivity and
fed on cases of crescents had been found to contain pigmented cells;
while these cells could not be seen in insects of the same kind which had
not been so fed. Just at this time I wrote to Manson, in a state of un-
bounded delight, that he might expect to know the full life-history of
Ihe parasites of malaria in the mosquito within a few weeks.

Next day however, I received telegraphic instruction from Government
ordering me to proceed forthwith to Kherwara in Rajputana — a place a
thousand miles distant!

14. Interruption; September iSpy — February i8g8. — It would be
^difficult for others to understand the effect of this cruel blow. Here in
Secunderabad I had numerous cases of malaria in my own hospital, and,
moreover, the men had been trained to submit to mosquito bites — a mat-
ter often of some difficulty with the superstitious natives of India. I had
also experienced assistants hired by myself for the work; and, above all,
the proper kind of mosquitoes, including their larva;, just found in abun-
dance. There is no doubt whatever that, had I been left at Secundera-
bad, I could easily have traced the whole Hfe history of the human pa-
rasites in dappled-winged mosquitoes within a few weeks. But at Kher-
wara I did not know what would happen. It was in the north; winter
was approaching; and I knew that mosquitoes would refuse to bite in

40

the cold. I failed even to guess the reason for this sudden transfer. The
astonishing discovery of the pigmented cells had been officially and full}'
reported to the Government through the chiefs of my own department;
malaria is the most important disease of India; and I thought that my
superiors were taking the greatest possible interest in researches which
touched so vital a subject — I thought that they would make every effort
to leave me undisturbed, if not to give me active help.

But the orders were peremptory and not to be discussed. Within
two days (26th September) I was on the week's journey to Kherwara..
I saw only one gleam of comfort. It was impossible that my chiefs, me-
dical men, would consent to interrupt my work at such a moment. There
must undoubtedly be a bad outbreak of malarial fever at Kherwara which
would throw great light on my subject.

When I arrived at the place however — a petty station with three or
four Europeans (whom I shall always remember for their kindness), and
part of a native regiment of Bhils, isolated in the midst of miles of wild
country far removed from civilization — I was told that there was no mal-
aria there; there had not been a case for months!

This then was my Elba — almost my lie du Diable; and I saw no
prospect of escaping from it for a year at least. After excusing myself
from accepting the appointment in Berar, I had indeed later asked to be
remembered for a permanent appointment to which I thought my long
service (more than sixteen years) and my work at Bangalore had at least
given me some claim. But this was only a temporary and insignificant
one, generally held by juniors ; and I do not know why the transfer was
made, unless possibly (though not certainly) for reasons connected with
the Afridi war. At all events it was made without reference to my re-
searches. I wrote officially to my superiors, begging to be allowed to re-
turn to Secunderabad to continue my work; but received only a reprimand
in consequence. There was no escape; but my pension was due to
me the following April, and I made up my mind to apply for it as
soon as the war was over, and to continue my researches as a private
person.

The cold weather came on apace, and at first it appeared to be ut-
terly impossible to work. There were no cases of malaria and scarcely
any mosquitoes. Much to my pleasure, however, I found a few dappled-
winged gnats, and observed again that their larvee lived in water on the
ground — namely in a pit and an old well — apparently almost as dor-

41

mant as the adults were. I kept a single one alive in a bottle for two
months without its developing.

Shortly after arrival at Kherwara I wrote down a brief account of
the finding of the pigmented cells in the third and fourth mosquitoes.
At the end of January the British Medical Journal containing my previous
paper on the cells [38], together with remarks by Manson, Bland Sutton,
and Thin, reached me. I therefore rewrote the beginning of my second
paper; and added a reference to some work which I had been able to do
with pigeons, and also a long discussion of Thin's remarks, in which I
showed that his position with regard to the pigmented cells was un-
tenable. The paper was published in February. I did not explicitly say
that the third dappled-winged mosquito had been bred from the larva in
captivity, because it was evident that this fact would be inferred from
the opening of the first paper of which the second was obviously a con-
tinuation. But I said that the grey mosquito in which pigmented cells
had been found was ^observed feeding on a patient», and that »I judged
for many reasons that it had been feeding occasionally on the same man
for several days > — showing clearly enough that this insect had not been
bred from the larva in captivity. The facts might have been put more
explicitly at the time; but they are apparent enough to any candid read-
er.* In the paper the order of the third and fourth mosquitoes is chang-
ed for purposes of description — the case of the grey mosquito being put
last because it was doubtful.

The work with pigeons just referred to was as follows. Being unable
to obtain cases of human malaria I turned to the malaria of birds which
had long been known to harbour parasites closely similar in appearance
and life-history to the malaria parasites of man. Both Manson and I had
long recognised the technical advantages of working with these organisms.
I immediately found the parasites of Labbe's genus Halteridinm in the
pigeons of Kherwara; but could not induce mosquitoes to bite the birds.
Observing, however, that they were infested by a species of blood- sucking
fly, I examined thirty of these, and some lice, fed on infected pigeons.
No pigmented cells were, however, found in them.

* When I wrote these papers I did not suspect that every line of mine, even in some
of my private letters, would be subjected to a minute and unscrupulous analysis in the hope
of finding discrepancies which would serve to discredit my observations. Every possible ar-
tifice has been used for this purpose by the very men who learnt all they knew from these
very publications.

42

At last when the weather became warmer in February several cases
of quartan fever occurred among the troops, probably relapses. The dap-
pled-winged mosquitoes still refused to bite; but I succeeded in feeding
a number of brindled mosquitoes of a peculiar brown species on the
cases. The results were again negative in thirty-four of these insects.

I was just about to apply for my pension when welcome news arriv-
ed. I had of course given full details of my sudden transfer to Manson;
and he had exerted himself to influence the government of India and the
Director General of the Indian Medical Service (then Surgeon General
Cleghorn) to put me on special duty to continue my researches. I had
urged the same thing upon the Director General; but, unfortunately as
it happened, suggested that one good place for the work would be Assam,
where an epidemic of kala-azar — a disease which Rogers had recently
reported to be malaria — had long been raging. However, I now received
a telegram stating that I had been placed on special duty to investigate
malaria and kala-azar in Calcutta and Assam for six months.* My five
months' imprisonment was at an end. I arrived in Calcutta on the 17th
February 1898; and was joined there by my family, with all my books
and notes which had been with them at Bangalore all this time.

15. Calcutta; Febi'uary — Aprils i8g8. The Theory Proved. — Now
in recompense for the tribulations of Kherwara, opened a glorious time,
during which the amazing story of malaria was unrolled little by little.
The great induction had given the clue; now, following the clue step by
5tep, we were to be led into regions where nature revealed herself won-
derful beyond the imagination of any of us. In the background was
something greater still — the possibility of saving human life on the large
scale.

I am happy to be able to begin this part of the narrative with a
brief account of the brilliant and important discovery of MacCallum. It
will be remembered that Manson had thought the motile filaments to be
flagellated spores; that I had studied them much without being able to
learn anything new about them except that they are certainly living or-
ganisms; and that when I finally found the pigmented cells I thought
that these were derived from the motile filaments, and had absorbed their
melanin from the haemoglobin in the stomach cavity of the insects. In
his letter of the nth August however, Manson sent me a paper by Si-

* Afterwards extended to one vear.

43

mond, suggesting that the similar motile filaments of certain Coccidia are
not of the nature of flagellated spores at all, but of the nature of sperms
[35]. How were these facts to be reconciled?

In a letter dated the 17th November 1897 Manson informed me that
a discovery had been made by W. G. MacCallum in America regarding
the motile filaments, showing independently that they are of the nature
•suggested by Simond's work. He did not send me the literature; and as
his letter reached me at Kherwara I could not then obtain it. Shortly
after my arrival at Calcutta however, I procured a copy of the Lancet
[36] which gave an abstract of MacCallum's work. The discovery was
as follows.

In 1897 Mac Galium undertook a study of the motile filaments. Work-
ing with the Halteridiuni of birds he noticed first that the gametocytes
seemed to be of two kinds, namely one kind which produced the motile
filaments, and another kind which did not do so. On watching two of
these cells, one of each kind in the same field of the microscope, he ob-
:served (July 1897) ^^^^ ^he filaments escaped from one as usual; that it
moved about actively for a time; and then approaching the other game-
tocyte actually entered it. Other observations of MacGallum and Opie,
made both on Halteridium and on the crescentic gametocytes of the aes-
tivo-autumnal parasite of man, confirmed this beautiful discover}\ The
fact, as previously shown by Sacharoff, that the filaments contain chro-
matin was now explained; and also the facts that they escape and move
about in the blood. They are, indeed, sperms which are emitted from
the one kind of gametocytes, the males, and which fertilise the other
kind, the females. Thus these minute parasites, among the lowest of
■creatures, have their sexes, and a form of sexual reproduction precisely
hke that of the highest animals.

More than this, MacGallum observed in the case of Halteridium of
the crow that the female cell, motionless before fertilisation, afterwards
becomes elongated and vigorous, and moves across the field in vitro.
This motile form had apparently long been seen by Danilewski and had
been called by him a vermicule*

So much for the motile filaments, but now what were the pigmented
-cells? Everyone seems to have thought that as soon as the flagellate

* I should certainly have observed these facts when I was making a special study of
the^ motile filaments in 1895 ^"^^ 1S96. I repeatedly saw them apparently attacking leuco-
cytes [42, page 14]. The reason why I found that only a percentage of crescents emit the
filaments in the mosquito's stomach is now explained — the remainder were females (section 11).

44

spores disappeared, so did Manson's theory. But it was not so. The
induction remained as strong as before; the locus of the phenomenon was
still in all probability the stomach-cavity of the mosquito. MacCallum's
work seems to have reached Manson shortly after my discovery of the
pigmented cells came to him. He connected the two groups of facts in
a moment. My pigmented cells loere the vermicides^ or fertilised female
cells, which had btirrotved into the insect' s tissues for the purpose of
undergoing further development there. This, and not my hypothesis
made before MacCallum's paper was known to me, explained the pre-
sence of pigment in the cells. He communicated his views to me in his
letter of the 7th February, and published them later [41].

Meanwhile, after another struggle, I was again in sight of the pig-
mented cells. On my arrival at Calcutta I found myself installed in the
convenient little laboratory which had been formerly used by Professor
D. D. Cunningham. There was a native assistant there; but I hired at
my own expense several others, especially a most intelligent Mahommedan
named Mahommed Bux, who after he had been trained showed great
enthusiam and gave me much assistance. To my delight I at once noted
several varieties of dappled-winged mosquitoes, besides man}^ kinds of
grey and brindled mosquitoes, actually within the laboratory, and found
the breeding-places of the latter just outside. Those of the dappled- winged
mosquitoes were detected a little later; and were again seen to be pools
of water on the ground. The next thing was to obtain cases of malaria;
but here I was met by an unexpected and most unforeseen misfortune.
The plague had been raging all this time in India; and on the Govern-
ment's trying to introduce Hafifkine's proph\'lactic inoculation in Calcutta
just before my arrival, serious riots, during which many of the Europeans
had felt themselves obliged to go about armed with revolvers, had occur-
red. The ignorant populace, thinking that the British were trying to
inoculate them with and not against plague, flew into paroxysms of terror
at the very sight of a European hakim (physician), while anything re-
motely resembling inoculation made them frantic. The physicians of the
Calcutta hospitals w^ere evidently very unwilling that I should use their
cases for my experiments under these circumstances; and, as I had no
hospital of my own as in Secunderabad and Bangalore, I was forced to
send my assistants into the bazaar (native parts of the city) in order to
try to induce patients to come to me on payment. Calcutta is not very
malarious, especially at that time of the year, and it was only on large

45

payment that several beggars with fever were induced to come to me;
but when I proposed to prick their fingers in order to examine their
iDlood they generally left their money, took up their crutches, and fled
without a word! This placed me in complete perplexity as to what to
do, until I remembered the malaria of birds. A number of crows, pi-
geons, weaver-birds, sparrows, and larks were then immediately procured,
and experiments commenced on them without delay.

The malarious parasites of birds are exceedingly closely related to
those of men, and together with these and the malaria parasites of bats
and monkeys form a group which is quite distinct from the intracorpus-
•cular protozoa of some mammalia, such as the Pyrosoma bigeminum of
<:attle, and of reptiles, such as Drepanidunn. The true malaria parasites
(namely the intracorpuscular protozoa of man, birds, bats, and monkeys)
are distinguished by their generally amoeboid character, by their posses-
sion of the characteristic black or brown pigment (melanin), and by an
identical life-history as regards the production and appearance of the
spores within the corpuscles, and of the motile filaments shortly after the
blood containing them is drawn from the host. The parasites of birds
■differ from those of man only in some very small morphological details;
and are so similar that in the earhest sub-classification of the group by
Grassi, one of the parasites of birds commonly called Proteosoma is plac-
■ed with two of the human species, the quartan and tertian, in one genus;
while the other parasite of birds, commonly called Halteridiuin^ is placed
in another genus together with the remaining parasite of man, that of the
pernicious, remittent, or sestivo-autumnal fevers. The latter part of Gras-
si's classification was wrong; and we now recognise that both the para-
sites of birds must be placed in one group with the quartan and tertian
parasites of man; while the third human species must be placed in a
group by itself, owing to the distinct shape of its gametocytes (crescents).
Thus zoologically, the avian species are actually more nearly related to
two of the human species than these are to the third human species.
Anyone who had actually studied all these parasites, moreover, would
Tiave little doubt that they would be found to possess practically identical
life-histories outside the vertebrate hosts, or at least life-histories which,
if not identical, would be closely similar. It did not of course follow
with certainty that the carrying agents of the avian parasites would be
the same as those of the human species; but we could safely assume
that they would be some kind of blood-sucking arthropod. At all events

46

it was certain that the discovery of the hfe-history of the avian parasites
would immediately open up that of the human organisms; while the prac-
tical difficulties of working with birds and infecting them would be less
than with men. In fact I should have been wise to have begun my re-
searches with birds in 1895. I therefore determined to employ birds at
once pending the subsidence of the plague-scare, when I purposed of
course to return to the human parasites; and there is no doubt that this
was the right course.

It was first advisable to see whether mosquitoes would not carry one
or both of the avian parasites. A number of crows and pigeons had been
found to contain Halteridmnr, but without waiting to examine the other
birds, I placed one crow, two pigeons, four larks and six sparrows in
several cages all within the same mosquito netting, and then in the eve-
ning released within the net a number of grey and brindled mosquitoes
bred from the larva in captivity. Next morning many of the grey mos-
quitoes were found gorged and were collected and kept for several days
according to my rules. On the 13th and 14th March I dissected them
one by one. When thirteen had been examined with negative results I
began to fear that I had committed myself to another tedious search for
the proper kind of host of the avian parasites. But fortune was kinder
on this occasion; the fourteenth mosquito had pigmented cells precisely-
similar to those which I had found in the dappled-winged mosquitoes fed
on patients with crescents.

Next I examined the larks and sparrows used in this experiment
together with the crows and pigeons, and found that they contained not
Halteridiuni but Proteosorna; so that it was doubtful from which kind of
parasite the pigmented cell had been developed. Consequently I now put
the birds with Halteridiuni in one net and those with Proteosorna in
another, and released within both nets numbers of grey mosquitoes bred
in the same bottle. Of thirty-four of these fed on the birds with Halte-
ridijini all were negative; but out of nine fed on the birds with Proteo-
sorna^ no less than five contained pigmented cells.

This result was obtained on the 20th March and practically proved
the mosquito theory of malaria. Out of hundreds of grey mosquitoes
previously examined none had contained pigmented cells except one which
had been caught feeding on a case of tertian (section 13), and one which
may have bitten one of the birds with Proteosorna in the experiment of
the 14th March. Now, however, no less than five out of nine fed on birds

47

with Proteosoma contained them. Mathematically therefore the probabili-
ties were enormous (amounting almost to certainty) in favour of the view
that the pigmented cells in this experiment had been derived from the
Proteosoma. The cells were in the tissues of the insect; the parasite must
therefore be able to make its way into and live in mosquitoes; precisely
similar cells had been found in mosquitoes fed on men with malaria — and
the chain of proof was complete.

But the fact that the pigmented cells in the mosquitoes are indeed
derived from the parasites in the birds was of such fundamental import-
ance that it required the most formal and rigid proof — especially as no
life-history of a protozoal organism able to transfer itself from one host
to another was then known to science.*

I therefore now commenced a long series of differential experiments
in order to establish the fact thoroughly. Grey mosquitoes bred from
the larva in captivity were fed (a) on birds with Proteosoma and (b) on
birds without Proteosoma^ and the results compared. The details will be
found in my Report [42]. Out of 245 grey mosquitoes fed on birds with
Proteosoma., 178, or 72 per cent, contained pigmented cells, while out of
249 of them fed on blood containing other parasites or no parasites, not
a single one contained them.

Another experiment was the following. Three sparrows were select-
ed, one with no parasites, one with a few Proteosoma., and one with many
Proteosoma. They were placed in separate nets, and numbers of grey
mosquitoes from the same breeding bottle were fed simultaneously but
separately on them. Ten mosquitoes fed on each bird were then examin-
ed, and the total number of pigmented cells in all of them were counted.
The results from a hasty enumeration made by myself were as follows.
No pigmented cells were found in the ten mosquitoes fed on the sparrow
without parasites; 292 in the ten mosquitoes fed on the sparrow with a
few Proteosojna; and 1009 in the ten fed on the one with many Proteo-
soma [42]. The preparations were sent to Manson, who made a more
careful enumeration and found o, 571, and 1084, pigmented cells in the
three sets of mosquitoes separately [41].

The fact then was proved, and the theory that the parasites of mal-
aria develop in mosquitoes was practically established. Meanwhile I had
been proceeding in the fascinating task of watching the progress of that

* The life-history of Pyrosoma in ticks is not even yet known ; and the transference of
trypanosomes by flies appears to be merely mechanical.

48

development. A number of grey mosquitoes would be fed on an infected
bird and would be dissected two, three, four days, and so on, afterward.
It was thus found that the pigmented cells grew rapidly in size until about
the eighth day, when they became so large as to be almost visible to
the naked eye. At this point they seemed to become mature; and it
could be seen that many of them burst within the insect; because mos-
quitoes which had been infected more than eight or nine days before dis-
section were found to contain not the mature pigmented cells, but only
their empty capsules. For the moment I could not ascertain what became
of their contents.

This part of the work led to an interesting observation which influ-
enced all subsequent researches om mosquito-borne disease. It will be
remembered that Manson had always thought that a few days after her
meal of blood the female mosquito laid her eggs and died; at this mo-
ment he considered both filaria; and malaria parasites escape into the
water from the insect [26]. I had accepted this view, but had frequently
observed that the insects do not die immediately after laying their eggs;
and now, as I watched the pigmented cells growing larger and larger
without apparently ripening, even five days after the insect was fed, it
occurred to me that we had been allowing our mosquitoes to die so early
owing to a very simple reason — we had omitted to feed them again! I
therefore fed my infected mosquitoes a second and a third time, and
more; and found that I could easily keep them alive for a month.* This
enabled me to work out the development of the malaria parasites com-
pletely; and also helped others subsequently to find a further stage in
the development of filarise, and to ascertain the mode of infection in ^-el-
low fever.

I did not succeed, and, indeed, scarcely attempted to find the host of
Halteridiinn. Nor was there time to work out the formation and beha-
viour of the ; vermiculesv in the stomach cavity of the mosquito — although
this could have been done very easily; but on one occasion I saw the
motile vermicule of crow's Halteridmm in a brindled mosquito.

Of course all this time anxious efforts had been made to obtain cases
of human malaria for experiment. Early in March I succeeded after much
difficulty in finding an old beggar with a few crescents willing to submit
to the dreaded operations; and I examined 41 grey mosquitoes and 15

* I refed them on healthy birds: but Bancroft subsequently found that they could be
kept alive for some time on bananas. ,

49

dark greenish dappled- winged mosquitoes which had been fed on him.
The first kind were tried merely as controls, and were of course nega-
tive; but, much to my surprise and disappointment, so were the latter.
I attributed the failure to the facts that the crescents were very scarce in
the patient, that the mosquitoes fed very sparingly, and that there was a
spell of very cold weather (for Calcutta) at the time. A few unsatisfac-
tory experiments with grey mosquitoes fed on a child with mild tertian
parasites also failed. In spite of all efforts no other cases could be pro-
cured.

A full list of all these experiments, beginning with my earliest work
in 1895, will be found in my Report written a few weeks later [42].

Recognising, of course, the inadequacy of my nomenclature for mos-
quitoes and the urgent necessity for employing the correct entomological
names for the various species used by me, and having failed to obtain
any literature on the subject, I now applied for assistance at the Indian
Museum in Calcutta; but I received a brief reply to the effect that the
savants there could give me no information on the subject. Once more
I had to depend on myself; and I therefore took special note of the dap-
pled-winged mosquitoes found near my laboratory. No less than four
species were detected — a large brown species, a large greenish one (with
which the experiments just described were made), a small black one, and
a small brown one. The first was named later by Giles from specimens
brought to England by me, and was called by him Anop]ieles rossi; and
from the studies of Stephens and Christophers made in Calcutta some
years subsequently it is almost certain that the second species \va.sA./u-
liginosus.

Numerous specimens of Proteosoma in grey mosquitoes were sent to
Manson on the 30th March.

By the middle of April I had overworked myself, and was obliged
to ask for ten days' leave to the Himalayan hill-station Darjeeling, where
I hoped for time to write my report in a cool climate. I had heard also
of several intensely malarious spots at the foot of the Darjeeling moun-
tains, and hoped to be able to carry on there the studies on human mal-
aria which were debarred in Calcutta, and at the same time to continue
my work on avian malaria. I therefore left Calcutta on the 17th April.

16. The Darjeeling Terai; April — June, i8g8. Efforts to Obtain
Assistance. — The results with Proteosoma were obviously so important

Researches on Malaria. 4

APR 2 0 1920 I -^ —

^ . 4/

\ that it wife necessary to give them to the world at once, in the hope that

^^■"---^-Hiljai^T^robservers would now be easily able to follow the work, and
also that I might obtain assistance in consequence of my success. Conse-
quently I devoted my time at Darjeeling to writing a report to my chief,
the Director General of the Indian Medical Service, on my latest work.
The report begins with a brief statement of my first discovery of the
pigmented cells,* followed by a list of the experiments, both positive and
negative, which I had made with a view to infecting mosquitoes with
human malaria. Then comes a detailed account of experiments and posi-
tive results with Proteosoma, followed by a minute description of the ne-
cessary technique, and of the appearance, position, and development of
the pigmented cells. Next I discuss several points, including the bearing
of MacCallum's work on mine. As I had brought my microscope and
some of my specimens with me, I was able to add to the report large
plates giving drawings of the pigmented cells up to the stage to which
they had as yet been traced. f The work was, however, hurriedly execu-
ted, as I had only a few days in which to write it. The pigmented cells
are called in it »proteosoma-coccidia», a term which has been criticised.
I thought at that time that the parasites of malaria really belong to the
Coccidiidse, the early stages of their life being passed in man and birds,
and the later stages (to which the name Coccidia might m.ore appropri-
ately be attached) in the mosquito; just as the early and later stages of
the sexual forms of C. oviforme occur respectively in the bile ducts and
the intestine of the rabbit. At the end of the report a description of
the grey and brindled mosquitoes with drawings is furnished by Mr. G.
C. Dudgeon, a gentleman who was acquainted with entomology; and the
report concludes with the words, »These observations prove the mosquito
theory of malaria as expounded by Dr. Patrick Manson. . . .»

The report after some delay, was dated 21st May, and was despatched
at once, with an urgent request that it might be published as soon as
possible. To my surprise I was informed that publication was not allowed
without the permission of the Secretary of State for India. This meant
writing to England and several months delay; but the report was printed
very soon and numerous copies were sent at the end of June to Manson
for private circulation among persons interested in malaria. In the mean-
time my success had been described in detail both to Laveran and Man-

* In the twelfth line the word Jordinary» is a slip of the pen for I'Others.
■j" These plates are reproduced at the end of this publication.

51

son in letters dated 22nd April — the letters being accompanied by a series
of seventeen more preparations; and, as my results could not be published
by myself, I now asked Manson to publish them for me.

Ik

''^/

I'ig. I.— From a preparation of mosquito's stomach dissected thirty
hours after the insect had fed od bird's blood containing proteu-
soraa. Tlie pigmented cells evidently lie bcweeii the longitudinal
muse lar libre3 which they have to some extent disassociated.

30 hrs. srddav.

■-J

6th day.

Fig. 2 —Development of the piji:'?: tod cell. The figure marked
"6th day" i:5 intended Ti rep.-cser.t what is evidently a capsule
from which its contents have escaped.

Figure _?. From paper by Manson, British Medical Journal, i8th
June T898, page 1577. After Ross's drawings.

On the 1 8th June Manson published an able paper on the subject.
The article commences with a resume of my original discovery of pig-
mented cells in dappled-winged mosquitoes fed on a human patient with
malaria, and gives the references to my papers describing the observation
[41]. It goes on to describe the new results with Proteosoma; giving

52

drawings of the pigmented cells up to the sixth day of development, and
a diagram showing the connection between MacCallum's observation and
my own; and it concludes with letters from Nuttall and Laveran accepting
my results. Laveran said, »It appears to me to be undoubted that the
elements discovered by Dr. R. Ross in the stomach of mosquitoes fed on
the blood of birds, the subjects of haemosporidiosis, are really parasites,
and that these parasites represent one of the phases of the evolution of
the haematozoa .... I have shown the preparations to M. Metchnikoff, who
shares my opinion. »*

This paper drew general attention to my w^ork, to which previously
little credence had been attached; and, as many of my preparations had
been sent to England and France, not only were those competent to form
an opinion enabled to judge of the truth of my statements, but those who
wished to follow my steps were now easily able to do so. In fact a most
amusing comedy now commenced, in which we witnessed the hasty efforts
of those who had been sceptics, not only to follow my steps but to per-
suade the world that their labours were original. During several years
since that date every observation of mine has been independently discovered
by various writers.

Recognising the vast significance of these preliminary results with
Proteosoina, and also feeling that it was quite beyond the power of one
man to complete as quickly as the interests of humanity demanded the
work which remained to be done, I now made strong efiforts to obtain
assistance. The help of a single medical man to collect mosquitoes and
cases of malaria for me would certainly have enabled me to reach the last
proofs in a month or two; and be it remembered, the mortality from
fever in India alone is said to amount to something like ten thousand
persons every day. When, however, I asked the Director General for the
services of one or more junior medical officers, I was told that none could
be spared at the time. As a matter of fact there are always many me-
dical officers in military employment in India, who can be spared if they
are urgently called for; and the truth is that the necessary trouble was
not taken. I then wrote to Manson begging him by all means in his
power to obtain assistance for me from England; and thought that the
Royal Society, which is subsided to a small amount by Government, might
afford to give it. The matter was considered; and it was finally agreed

* Owing to a misapprehension, this paper erroneously states that Halteridium also had
been cultivated.

53

to appoint, with the help of the Colonial Office, a commission of three
gentlemen to investigate malaria. Two of these were sent in the autumn
to study the subject — in Italy; and, after much difficulty, the third was
allowed to come to me. He arrived at Christmas with orders to stay for
two months — not to help me but to verify my statements!

That was all the help I received. The excuse is that my work had
not been confirmed. But it had been accepted by Laveran, Manson,
Metchnikoff, and Nuttall, who at least knew the subject. Was not this
enough to justify the expenditure of a few hundred pounds in so great a
cause? I mention these facts because it was largely this failure to obtain
assistance which drove me from India some months later; which delayed
the completion of my work for more than a year, and which postponed
the adoption of an energetic prophylaxis in India until the present. Not
mine the fault: the truth is that for some inexplicable reason men will
never recognise the transcendent importance of investigation into the causes
of those great diseases which destroy them.

The rest of my time in this district was spent in making attempts
to find a suitable place in the intensely malarious areas at the foot of the
mountains for researches on human malaria. This alone was a matter of
no little difficulty, as the locality was new to me and I could obtain no
accurate information regarding the disease. I worked especially at a place
called Punkabari, situated a few hundred feet above the plain. A hospital
and plantation existed here, and there was a large village some miles away
on the plain. But the results were not gratifying; few dappled-winged
mosquitoes could be found, as the rainy season had not yet commenced;
while to my grief I discovered that the plague-scare was, if anything,
stronger here than in Calcutta. So terrified were the natives, that on one
occasion, when one of my men shot a sparrow for me in the village, all
the coolies in the neighbourhood ran away for miles into the jungles,
costing the planters much money and trouble before they could be in-
duced to return. In fact I was given to understand that scientific in-
vestigations were not required there at the moment! Indeed it soon
became apparent that I was only wasting much valuable time; and I
consequently determined to complete my researches on Proteosoma at
Calcutta without further delay.

17. Calcutta; June — August, i8g8. The Route of Infection. On my
return to Calcutta (4th June) I found it still quite impossible to obtain

54

cases of human malaria for my work, and therefore proceeded at once
with the Hfe history of Proteosoma. The most wonderful of all the phases
of this history was now to be revealed. I had traced the development of
the pigmented cells up to their maturity and subsequent rupture and dis-
charge of their contents into the body-cavity of the grey mosquitoes. I
could not see at the moment what happened to these contents; yet upon
this point depended the vastly important question of the route of infec-
sion in malaria. But, when I had broken off my work a few weeks
previously, the contents had appeared to consist of little more than a
pure fluid.

Hitherto my mosquitoes had been dissected in water or a weak solution
of salt, and I had had no time for methodical staining. A strong salt solution
was now used and the secret was revealed. The contents of the mature
pigmented cells did not consist of clear fluid, but of a multitude of delicate
thread-like bodies, which, on the rupture of the parent cell, were poured
into the body-cavity of the insect, and which were evidently spores.

jj

T^))y

li{<^{^\

Figure 4. Sketch of thread-like bodies (sporozoids or blasts)

escaping from mature ruptured pigmented cell (zygote). From

letter of Ross to Laveran, dated i8th July 1898.

What happened now to these spores in view of the theories mentioned
in section 12? Did they escape into the water according to Manson's
ideas; or were they voided by the intestine according to mine; or did
they in some mysterious manner work their way into healthy persons
during puncture, according to the theories of King and Bignami and later
of myself? But the staff of theory was no longer necessary ; plain research
would suffice.

Here there was another sharp but short struggle. I saw that the
thread-like bodies, although apparently without motion themselves, were
soon scattered by the insect's circulation all through its body; but beyond

55

this I could not follow them for some time, in spite of the most assiduous
endeavours. They seemed to have been created without object.

On the 2nd July however, I found in the thorax of a mosquito a
large cell which, surprising to state, contained within it several of the
thread-like bodies. They were able then to work their way into cells;
but what was the cell? On the 4th July, while working upon another
mosquito, I found that the thread-like bodies seemed to become more and
more numerous towards a point in the thorax — as if they were converging
toward some destination. At that point there were numerous cells such
as I had seen on the 2nd July. They were attached to a duct and were

Figure 5. Thread-like bodies (sporozoids) in cells of salivary gland of mosquito.
From letter of Ross to Laveran, dated. i8th July 1898.

all contained within the same capsule — they constituted in fact some
kind of gland. In all these cells there were hundreds of the thread-Uke
bodies, floating loosely at all angles to each other like fish in globes of
glass. Close by was another lobe of the gland similarly full of the spores.
I was at the summit but not on it. I did not know what the gland was.
I knew the appearance of the cells it is true, but in spite of my thousand
and more dissections I had by no means acquired a full knowledge of
the macroscopical anatomy. I found it by no means easy to meet with
the gland again. On the 8th July the mystery was solved. The gland lay
in the neck and upper thorax — the throat — of the mosquito. It con-
sisted of three lobes on each side. The ducts of each lobe unite together
Hke the midrifs of a trefoil. The duct so formed runs forward and meets
the similar duct of the other side under the chin — so to speak — of
the mosquito. The common duct advances still further and enters through
the round base of the central stylet or stabbing weapon of the mosqui-
to's proboscis. It was easy now to recognise the nature of the gland;

56

it was the salivary gland^ which secretes the irritating fluid which
the mosquito injects in the wound made by her in the skin, perhaps to
dilate the vessels, perhaps to prevent speedy coagulation of the blood.*
The exact route of infection of this great disease, which annually slays
its millions of human beings and keeps whole continents in darkness, was
revealed. These minute spores enter the salivary gland of the mosquito
and pass with its poisonous saliva directly into the blood of men. Never
in, our dreams had we imagined so wonderful a tale as this.

But still all this was inference only; the last proof was demanded.
If the infection can be given in this way, give it. I had long possessed
in the laboratory five old birds — four sparrows and one weaver bird —

which had been kept there for my
»controb> experiments, because they
had never been found to contain
Protesoma^ even after several exa-
minations. On the 25th June, as
soon as I began to suspect the
destination of the thread-ilke bodies,
these birds were all examined again,
and were found to be still quite
healthy. On that and the follow-
ing nights, a large number of grey
mosquitoes which had been long
previously fed upon infected birds and many of which had been found
to contain the thread-like bodies in their salivary glands, were re-
leased within a mosquito net in which the five healthy birds were placed.
On the following mornings I satisfied myself that the infected mosquitoes
had gorged themselves freely on the birds; and then, fascinated by the
study of the parasites in the salivary glands of mosquitoes, I forgot all
about even this important experiment.

Now only a small percentage of birds in Calcutta are infected with
Proteosoma. Out of in wild sparrows examined by me I found the
parasites only in 15, or 13.5 per cent. Moreover, even in infected birds,
the parasites were scarce, seldom more than one being found in each field
of the microscope. On the 9th July I suddenly remembered my experi-

Figtire 6. Salivary gland of mosquito. Froi
letter of Ross to Manson, dated 6th July i8i

* This gland had been discovered in 1888 by Macloskie [5], but I did not know it at
the time and still had received no literature on the subject.

57

ment and examined the previously healthy birds. All of them without
exception were now found swarming with Proteosoma, as many as twenty
or even more being found in each field.

But not content even with this I repeated the experiment over and
over again; and within the next few weeks I succeeded in infecting 22 out
of 28 healthy sparrows (79 per cent), and also a crow and four weaver
birds, and, moreover, gave a more copious infection to four sparrows which
previously contained only a few parasites. At the same time I kept as
controls a number of healthy birds in mosquito-nets, safe from the bites
of mosquitoes, and found that none of them became infected (wMth one
exception probably due to an error).

Figure 7. Thread-like bodies (rods, sporozoids) in salivatory glands

of mosquito. Published from Ross's drawing by Manson, British

Medical Journal, 24th September 1898, page 852.

Manson, to whom I had sent full details, told me that he would ex-
pound all my results, with demonstrations of my specimens, at the meet-
ing of the British Medical Association to be held at Edinburgh at the end
of July. I now announced the successful infection of birds to him by a
telegram, which reached him just as he was setting out (though ill at the
time) for the meeting; and he was therefore able to communicate the
complete life-history of the parasites in his address.

His exposition as Dr. Charles said, ^created quite a furore*, and was
quickly made known everywhere. His papers was published on the 24th
September [43] and gave a full account of the subject up to the infection
of healthy birds, together with several drawings of the thread-like bodies,
both free and in the salivary glands, taken from my letters.

58

It was interesting during these researches to watch the gradual
invasion of the birds by the parasites. From five to eight days after
they were bitten by infected mosquitoes no parasites could be found
in their blood; then a few appeared; then many; and at the last
large numbers. The first five birds all died, and so did some of the others;
and their liver was found to be full of the characteristic pigment of mal-
aria. But many recovered, the parasites gradually decreasing in number.

At the same time I was temporally not a little delayed by finding
inside the mature pigmented cells certain large brown or black bodies
which I provisionally thought might be connected with their life-history.
As proved by the researches just described, malaria could be carried by
mosquitoes from the sick to the healthy, but as we know malaria clings
intensely to location. It therefore seemed not at all unlikely that these
black bodies, occurring as they did actually within the pigmented cells,
might be of the nature of sporocysts meant in some way to infect other
mosquitoes — so that the infection might not only be carried from man
to man by the mosquito, but from mosquito to mosquito; or they might
be meant to infect man, as Manson had thought, through the water. It was
of course necessary, for sound science, to examine these bodies; and I
therefore tried to infect both birds and the larvae of mosquitoes by feed-
ing them on insects containing these black spores; but the results were
negative. Subsequently I saw reason to doubt whether the black spores
really had any connection with the parasites (section 20).

But there was little time for such researches, necessary as they seemed
at the moment. Although there could be no doubt that the human
parasites have the same history as Proteosoma, still it was a necessary
formality to complete the partial demonstration of this fact which had
been already attained, if only to persuade Government to take active
measures against the disease; and I was at last free to undertake the work.
But now precisely occurred my last and most annoying interruption.

Before coming to this, however, let us consider the results which had
already been attained and which have been the basis of nearly all that
has been subsequently done.

(i). The general life-history of Proteosonia in the grey mosquito
and the mode of infection being now ascertained, we could foretell to a
practical certainty that the life-history and mode of infection of all the
other parasites of the same group, including the human ones, would be
closely similar in all their stages; that is, that if they differed at all, they

59

would dififer only in small details. The result of this was that if anyone
wished to trace the life-history of any of these organisms in a second
host he would now find the task an extremely easy one, because (a) he
would know exactly the appearance of the parasite he was in search of,
and (b) he would know exactly in what part of the anatomy of the second
host to look for it. And if he wished to ascertain whether a given animal
was or was not the second host of the parasite, he could easily make
sure of the fact by ascertaining whether or not it harboured the described
parasites, after feeding and dissection by the methods laid down by me.

It is in fact solely by this means that we have been able to demonstrate
the proper hosts of the human parasites in many parts of the world.

(2). More than this, the pigmented cells of the aestivo-autumnal
parasite of man had been demonstrated to be exactly similar to those of
Proteosoma on the second, fourth, and fifth days after infection of the
mosquito; and the hosts of this important organism were shown to be at
least two species of a special genus which could be recognised by its
possessing spotted wings and boat-shaped eggs (section 23); and were
clearly shown not to be my grey and brindled mosquitoes, the former of
which had been described sufficiently for recognition [39 and 42].

(3). The important law that not all species of mosquitoes can har-
bour a given parasite of this group had been established, both with regard
to the aestivo-autumnal parasite and Proteosoma and Halteridhiin; and
several important facts regarding mosquitoes had slowly become evident
to me — but were not published until later.

(4). Lastly full directions of technique had been given in my report
[42]. These consisted of numerous essential details, acquired during several
years' experience, regarding dissection and feeding, etc. — without a know-
ledge of which the observer would be very likely to go wrong (as for
instance by attempting to section his mosquitoes for searching for the
parasites, and omitting to feed them regularly and change their soiled
habitations for clean ones).

On the other hand my researches had given little or no information
about the quartan and tertian parasites — except of course the all import-
ant analogy with Proteosoma. The observation of the grey mosquito
caught feeding on the case of tertian was doubtful (section 14). Moreover
they had not directly and absolutely demonstrated the final stages even
of the aestivo-autumnal parasites in the dappled-winged mosquitoes, nor
the mode of infection. But nevertheless they had reduced the demonstra-

6o

sions still required to an easy formality which was within the capacity of
any tyro with sufficient material and a microscope.

I am sorry to have to write such a summary of my work as this
one; but it is rendered necessary by those who, during the long inter-
ruption of my labours which now followed, were able to work out some
details of the subject before me, and who have wished to conceal the as-
sured fact that their efforts were simply a repetition and imitation of mine.
It should be pointed out that, by a generally recognised zoological rule,
the discovery of the life-history of Proteosoma in mosquitoes covers that
of other members of the same group of organisms, which have precisely
the same development. By that rule, the right of priority in discovery
belongs to him who first works out the life-history of one species of
a group of animals; not to those who merely perform the easy task
of extending the known facts to other species. Discovery is discovery;
the determination of parallel facts, the filling in of details, the publi-
cation of pretty illustrations, and the furnishing of formal proofs of
matters which are already certain, are useful — but do not constitute
discovery.

My infection experiments on birds were completed early in August,,
and, as will be related presently, I was now no longer able to defer my
work on kala-azar. Consequently I was obliged to leave Calcutta on the
13th August for my new duties — much exhausted by work and heat in
the plains. Before doing so, however, I released my host of little feathered
prisoners, which had unwillingly been of such assistance in the investigation.

It should be mentioned that from the first discovery of the thread-
like bodies I had wondered whether they have any other destination be-
sides the salivary gland. The eggs were especially suspected, but the
results of investigation were negative. I therefore now concluded that
malaria is communicated only by the bites of insects.

18. Darjeeling District; August — September, i8g8. Kala-diikJi. —
It was mentioned at the end of section 14 that I myself had proposed to
Government that kala-asar should be included in the programme of my
year's special duty; because I then hoped that this disease might shed
light upon the mosquito theory; but now when the theory was established
and it was necessary to press on with the study of the human malaria, I
wished to escape this additional duty, as I dreaded lest it should involve
me in much pathological work which would interfere with the principal

6i

line of research. I hinted as much to the Director General, but was told
that he expected me to adhere to the programme. The disease was
■exciting much comment because it was new and was taking some thousand
lives annually in Assam; but it was forgotten that malaria, though it is not
new, takes some millions of lives annually in India alone!

Harold Brown had recently studied a disease which existed at the
foot of the Darjeeling mountains, and which was called kala-dukh (black
sickness) and was evidently closely allied to kala-azar (black-fever). Con-
l sequently I obtained permission to investigate this disorder first, partly
because an opportunity might be afforded me of making some further
studies at the same time on malaria in my old haunts at Punkabari. Fix-
ing my head quarters at Kurseong in the hills on the road to Darjeeling,
I made numerous visits to this locality, but was dogged by ill-luck. The
plague-scare, though waning, was still present; and difficulties of transport
impeded the work. On the 25th August I arrived at Naxalbari, an intense-
ly malarious plantation and village on the plain beyond the foot of the
hills, and found swarms of small and large dappled-winged mosquitoes
(probably Anopheles listo7ii and A. rossi). There was no time to make
formal experiments, and the people would not have allowed them; but I
examined some dozens of these mosquitoes caught in the houses of infected
persons, both for the pigmented cells and the thread-like bodies; but with-
out success,* Nearly all my time was however taken up in patholgical
enquiries on kala-ditkh — as I feared would be the case. But now it
was no longer possible to postpone the evil hour without dereliction of
duty, and I was obliged to set out on the long journey to Assam.

19. Assam; September — November, i8g8. Kala-Azar. — I arrived
at Nowgong, the centre of the epidemic of kala-azar, on the 13th Sep-
tember. It was at once obvious that my worst fears were well-founded,
and that I would be plunged for months into a difficult pathological pro-
blem and a long pathological report. But the work was not without in-
terest, and I may be pardoned for touching upon it briefly. The disease
had been first noticed by McNaught in 1882. A few years later Govern-
ment sent Giles to investigate it; and Giles, who probably did not come
much in contact with the real disease, seemed to have been considerably

* How unfortunate I was in this respect may be gathered from the papers of Stephens
and Christophers [71] who later found a large percentage of these mosquitoes infected in this
very district.

62

misled, and in a report (which was nevertheless a very able one) pronounced
the malady to be ankylostomiasis [771. Many of the practitioners in
the locality were not satisfied however, and in 1896 Government sent
Rogers to make a further report. Rogers certainly saw the real disease
and concluded that it was a virulent form of malaria [78]. As it was
evidently communicable, this implied that he held malarial fever to be
communicable — a thing which no one would believe at that time; but
he maintained his opinions with great courage and success. I was now
sent in order, if possible, to decide the question; and as my researches
had shown that contrary to accepted views malaria must be communicable
from the sick to the healthy, Rogers position was justified. But the exact
nature of kala-azar still required definition; and, as I was called upon to
judge between opposite opinions, I was forced into a tedious enquiry —
though it was my immediate personal impression that the disease is malaria.
Mixed with the cases of kala-azar there were numerous cases ot
ordinary malaria; and I found that the local practitioners could not dis-
tinguish which was which until the cases became exceedingly severe, when
they were declared to be kala-azar. This generally happened only in the
later stages of the cases — so that in fact kala-azar seemed to be simply
another name for a very severe and frequently fatal form of malarial
cochexia. As, moreover, many of the patients had ankylostomes, those who
are familiar with the subject will understand that my task was indeed a
complex one. The plague-scare not having penetrated here, I attacked
the problem by examining the blood of all the cases, both of malaria and
of kala-azar. My results showed that while the parasites were easily
found in the early cases, they became more and more scarce as the dis-
ease advanced; until, in the old typical cases of malarial cachexia and
kala-azar neither parasites nor pigment were to be found, even in blood
taken from the spleen. I inferred then that kala-azar is probably only
malaria, though it was possible that some secondary infection might ac-
count for the gravity of the cases. I also inferred — what no one would
accept before then — that the spontaneous disappearance of the parasites
must be due to the gradual establishment of immunity; and that the low
fever present in these old cases was due, not to the parasites, but to some
secondary intoxication from the greatly enlarged liver and spleen. And
the same theories seemed to me to apply to kala-diikh.*

* It has just become highly probable that these diseases are due to a new parasite re-
cently discovered by Leishman and Donovan.

63

This investigation required repeated examination of the blood of all
the cases which I could procure in the town; and, being made at high
pressure, involved another extreme strain on the eyesight. Nevertheless
I examined several batches of dappled- winged mosquitoes fed on cases
with parasites, but the insects selected for the work were like some of
those abounding at Calcutta, namely Anopheles rossi. All proved nega-
tive. My disappointment was considerable, but I was not satisfied that
the feedings, which were left to assistants, were properly done. Many of
the same insects caught in the houses of patients were also negative. By
the aid of my assistants, however, many fresh examples of the law that
the dappled- winged mosquitoes breed in pools of water on the ground
were obtained.

During my stay at Nowgong I wrote a short report, dated the nth
October, on the infection of birds by the bites of mosquitoes [46]. This
was not published until some months later; but of course the principal
facts had long previously been published by Manson [43].

At the conclusion of my work on Kala-azar I returned, now utterly
exhausted, to Calcutta.

20. Calcutta: November i8g8 — February i8gg. The Work Con-
firmed. — Arriving at Calcutta on the 19th November I set to work to pick up
the threads of my work on Proteosoma; to obtain cases of human malaria
(the plague-scare having abated); and to write my report on kala-azar —
this being a tedious business requiring a full discussion of many intricate
details. But my health had now suffered greatly from the continuous exertion
made under very trying circumstances; and I felt scarcely able to complete
even my report. The labour, the disappointments, even the successes, of the
long and anxious investigations of a single subject had been too much for me.

The cold and dry weather had now commenced in Calcutta; and the
result was that the malaria parasites had become much more difficult to
find, either in men or in birds. Added to this, as I had no room in my
laboratory which could be warmed by a wood fire (a gas-stove injured the
insects), the mosquitoes could scarcely be persuaded to bite. And when
they did so, it was observed that the parasites developed in them much
more slowly than in hot weather.

Moreover, all this time I had failed to obtain any assistance in India,
and saw no prospect of obtaining any. I had been told indeed by Manson

64_

that Dr. Daniels was to arrive shortly; but he was being sent, not really
to assist me, but to enquire into the correctness of my statements; and
was to remain with me only for a month or two. The only persons who
had hitherto taken sufficient interest in my proceedings even to look at
my preparations (I mean from the beginning of my work in 1895) had
been Drs. Smyth, Maynard, Dyson, and Cooke; and it was clear that no
one really credited my results. Even the Director General, who was then
in Calcutta, would not visit my laboratory. It was the case of Galileo
and the satellites of Jupiter over again!

I was, however, much cheered by the arrival of Dr. Rivenburg of the
American Mission, who, hearing of my work, came all the way from a-
distant part of Assam with his wife and children at his own expense to
assist me. He had been previously quite unknown to me; and I shall
never forget his disinterested action and the help he gave me.

I had also been delighted to hear from Manson that the work was
now being taken up by Koch and the Italians. My papers had been publish-
ed as described; copies of my Proteosoma Report [42] had been sent
to many persons interested in malaria. On the 8th November Manson
wrote again informing me that he had just despatched some of my pre-
parations to Rome, namely to Bignami and Charles.

I did not become acquainted with the admirable work of Koch until
later (section 23); but the efforts of Bignami and Grassi were now commu-
nicated to me in a series of interesting and well instructed letters by Dr.
Edmonston Charles — a gentleman then staying in Rome, but whom I
had never met, nor corresponded with before. From these and their own
papers it was clear that the Italian writers had been inspired by my work
and had been desperately endeavouring to follow it; that they had detected
the genus {^Anopheles) of my dappled-winged mosquitoes, and, after hav-
ing seen my preparations, had succeeded at the end of November in find-
ing my pigmented cells in an Italian species of this genus caught in
infected houses. Bignami also claimed to have infected healthy persons
by mosquitoes obtained in this manner; and, by a lucky stroke, one of the
persons bitten by some A. claviger was found to have acquired the mild
tertian parasite.

I shall return to this subject when describing the confirmations of my
work. The efforts of Bignami and Grassi were, however, obviously hasty
and unreliable; while their writings were historically most inaccurate.
They therefore did not impress me, and exercised no influence whatever

65

in the completion of my own labours. For several years afterwards, how-
ever, nearly all my work was credited to these writers.

One interesting fact, however, I learnt from the Italians through Charles,
namely that my grey mosquitoes belonged to the genus Culex and my
dappled-winged mosquitoes to the genus Anopheles. Manson appears to
have sent some of the former to Grassi, and, according to Charles's letters
of the 19th November, he thought they were Culex pipiens — as a matter
of fact they were C. fatigans. Also from Charles's letter of the 25th
November and from their own publications [48, 51], it was clear that the
Italians thought that my dappled-winged mosquitoes were Anopheles cla-
viger'., and in his letter of the 6th January it was stated that Grassi con-
sidered some dappled-winged mosquitoes which I sent him were A. pictus
— they were really A. rossi, Giles. I satisfied myself more fully next
year at the British Museum regarding the zoological names of the mos-
quitoes studied by me. The Italians had no difficulties in these respects
as they had Ficalbi's works on gnats to guide them. They received some
more of my preparations, through Dr. Charles, early in January 1899.

On the 22nd December Dr. Daniels, of the Medical Service of British
Guiana, arrived. Though somewhat sceptical at first, he was soon con-
vinced after seeing my preparations and repeating the experiments with
care; and he fully confirmed my work in a paper, which however was
not published by the Royal Society until much later [71]. I am much
indebted to him for his assistance and advice in connection with my re-
port on kala-azar — no one has a profounder knowledge or a larger
experience of malaria. In the time remaining to us we attempted several
series of experiments on human malaria (cases now being more obtainable),
mostly with the large brown dappled-winged mosquito of Calcutta with
which I had made most of my few but negative experiments recently in
Calcutta and Assam. These too proved to be negative. We ascribed
our failure to some mistake; but the cause was ill-fortune. The mosquitoes
were chiefly Anopheles rossi which in Bengal certainly do not easily take
malaria.*

* It was, of course, the cold weather in Calcutta, but Daniels and I used incubators.
Stephens and Christophers later obtained positive results in Nagpur with A. rossi, but only
by very careful artificial regulation of the temperature [yi]-

It should be clearly understood that all my experiments on human malaria since 1897
were made under very unfavourable circumstances, and that I never considered my negative
results to be at all final.

Researches on Malaria. c

66

Several distinguished visitors came to us at this time — F. Plehn,
A. E. Wright, and A. Rufifer. They all accepted our work, except the
»black spores* mentioned in section 17. Nevertheless Daniels and I did
not think it right to abandon them without clear evidence; but when,
later, I found closely similar bodies in large numbers in mosquitoes which
had not been infected at all, my faith was shaken; and it was disturbed
still more when I failed to find them in the infected mosquitoes of Sierra
Leone. My doubts were mentioned in the concluding sentences of my
report on kala-azar.

That report was finished on the 30th January [79], and contains eighty-
one closely printed folio pages. My years' special duty was now almost
finished; but I could obtain no definite assurance from my chief that I
was to be retained on the same duty for an extended period. Yet the
matter was vital to me. Nearly all the money at my disposal had been
spent in consequence of these researches, chiefly because of the expenses
connected with the constant changes of station to which my family and

At this time I informed Daniels of some other details; and, in view of a controversy
which arose later, he has kindly testified to this fact in the following letter: —

October 8th 1900.
Dear Ross,
I shall have great pleasure in testifying to the following facts: —

(i). Shortly after my arrival in Calcutta in December, 1898, you showed me living
specimens of your >grey», 2>brindled», and sdappled-winged* mosquitoes.

You pointed out to me the attitude assumed by the last, the position of its larvse in
water, and the peculiarities of its eggs.

Since then I have learnt that these are characteristics of the genus Anopheles.
You contrasted these with the eggs and larvas of other mosquitoes, which I now know
to belong to the genus Culex.

(2). You showed me two species of s dappled- winged* mosquitoes, and I sent specimens
of them to the British Museum where they now are. They have been descril^ed by Major
Giles, I. M. S., as Anopheles.

You also showed me a specimen of a stomach of a mosquito with what are now known
as »zygotes2>. This you stated was the stomach of a » dappled- winged* mosquito, similar to
those you had shown me, which had been fed on a patient with crescents in 1897.

(3). I may add that the development of Proteosoma, as demonstrated to me by you in
the J'greys mosquito, is essentially the same as the development of >crescents2> in Anopheles as
observed by me in British Central Africa.

I am,
Yours very truly,
C. W. Daniels.
P. S. As regards breeding-places, you informed me that the »dappled-winged» mosquitoes
breed in puddles, the i'brindled» mosquitoes generally in flower-pots, and the »greyj mosquitoes
in tanks, ditches, etc.

C. W. Daniels.
Major R. Ross.

6;

myself had been subject; and if I were now compelled to return to Se-
cunderabad, I should not be able later to pay for my passage to England.
Moreover, both Daniels and Rivenburg were now leaving me, and it was
evidently foolish to expect any further assistance in India — much more
that of a trained entomologist, which I especially required for the completion
on my work on human malaria. I therefore determined to leave India
forthwith and to return to England, trusting to fortune to give me an
opportunity for finishing the investigation in a manner which I thought
suitable. I mention these personal details as I have been blamed for
leaving India at that moment.

Before doing so, I urged upon Government the importance of taking
active measures for the prevention of malaria in accordance with my ob-
servations. Besides advising the strict use of mosquito-nets for a personal
prophylaxis, I urged especially a campaign against mosquitoes as the best
measure for towns and cantonments — particularly against the dappled-
winged mosquitoes, which I said breed principally in water on the ground.
My letter was published later [55], and I hope that the advice will soon
begin to be taken.

I had also written a brief abstract of my work dated the 31st De-
cember 1898. This was presented by Laveran to the Academie de Mede-
cine on the 24th January 1899, and was published soon afterwards [54].
In this paper my obligations to Manson and Laveran were acknowledged,
I hope, in the full manner which honourable science demands. I wrote: —

«Pour eviter tout commentaire errone, qu'il me soit permis de
declarer ici que mes travaux ont ete entierement diriges par Manson,
et que j'ai eu I'assistance de ses conseils et de son influence a
toute occasion; je dois aussi remercier le Dr. Laveran de m'avoir
envoye ses avis si autorises. Quand, en mai dernier, je lui en-
voyai des specimens de mes corps pigmentes du moustique, il
reconnut immediatement la vraie nature de ces elements. »

And I added in conclusion,

«Je considere comme probable que la malaria est communiquee
a I'homme uniquement par les morsures des moustiques et peut-
etre d'autres insectes.»

21. England; March — Jiily i8gg. Foundation of the Liverpool School
of Tropical Medicine. — On the voyage to England (February 1899) I

68

had full time to consider the present condition of our knowledge about
malaria, especially in relation to the all important subject of prevention.
It was almost certain that infection is caused solely by the bites of in-
sects — but of what insects only? My long negative work had almost
proved that the commonest Indian mosquitoes, the grey and brindled
genera, do not carry aestivo-autumnal infection, at least. On the other
hand, it was certain that two species of dappled-winged mosquitoes in
Secunderabad, and one species in Rome, do carry it; while, if Bignami's
observation was to be trusted, the last species carries also the mild tertian
infection. But Secunderabad and Rome are not the whole world; even in
Bengal, Daniels and I had not succeeded in infecting dappled-winged mos-
quitoes. The question as to which species do or do not carry malaria
might prove to be a very complex one, not to be solved only by a few
local experiences; there are probably hundreds of species of mosquitoes
in the world, each of which would have to be tested unless we could find
some good reason for limiting the enquiry. I therefore sought for some
such reason, and found one. For centuries it had been known that mal-
aria is connected with stagnant water on the ground — not with water
in the pots, tubs, and tanks which abound close to all habitations, but
with marshes and pools on the surface of the earth. Again malaria was
known to increase every year at the rainy season, and subsoil- drainage
was known to mitigate if not remove the disease. Hence it was exU'eniely
probable that the insects which carry malaria breed only, or chiefly, in
terrestial zvater. For years we had assumed that the disease is caused
by organisms which spring from marshes. We had been partially right,
but not wholly right; it is not the infective but the infecting organism
which springs from the marsh — not the germ but the carrier of the germ.
Now, referring to mosquitoes alone, which varieties of these insects breed
only or chiefly in terrestial waters? I remembered my frequent observa-
tions on this point (sections 14, 15, and 19). The grey and brindled
mosquitoes breed chiefly in tubs and pots in India; but the dappled-winged
mosquitoes breed in pools on the ground. Now it was only these last
which, hitherto, had certainly been connected experimentally with malaria.*
What a weapon for good was now placed in our hands! Hitherto
when we wished to remove malaria we were obliged to drain a whole
area, recognising only that all terrestial waters seemed to be dangerous.

* This reasoning was by no means obvious or even known until after our work at Sierra
Leone. In temperate climates grey mosquitoes {Culex) also breed often in terrestial water.

69

Now we should be able to go to a place and to point out the actual pools
which cause the disease, by showing that they contain the larvae of the
culpable insects. The expense of dealing only with these would be
much less.

Shortly after my arrival in England in March I learnt something about
the zoological classification of mosquitoes from Mr. E. E. Austen of the
British Museum. I found that, as I had already partially learnt through
Charles, my dappled-winged mosquitoes were those of Meigen's genus
Anopheles and that both my grey and brindled mosquitoes belonged to
the same genus, namely Citlex. I was dissatisfied with this because it
seemed to me certain that they were of different genera. Recently Theo-
bald in his fine work on mosquitoes [75] has separated them, placing the
brindled mosquitoes in the genus Stegoniyia^ and reserving the name
Culex for the grey mosquitoes. Later, Giles determined that the grey
mosquitoes which carry Proteosoina are Culex fatigans^ and called
the large negative dappled-winged mosquitoes of Calcutta AnopJieles
rossi. *

Meanwhile Manson had been urging his great scheme of creating
special schools for the teaching of tropical medicine, and had now received
the support of Mr. Chamberlain. In Liverpool, Sir Alfred Jones, sup-
ported by Professer Boyce of University College, Mr. Adamson, Chairman
of the Royal Southern Hospital, and many other gentlemen, had warmly
taken up the scheme, and now appointed me the first lecturer of the
Liverpool School of Tropical Medicine. I therefore found myself no
longer an isolated worker, but a member of a company determined to
advance the interests of life and health in the tropics. And it was an
auspicious moment; for the great weapon which had just been forged for
the prevention of the most important of tropical diseases needed strong
hands to lift and wield it.

Almost my first care on returning to England was to consult eminent
zoologists regarding the proper nomenclature for use in connection with
the developmental stages of the parasites in mosquitoes. With the aid of
Professor Herdman I published a paper on the subject [59], in which,
abandoning the hasty provisional nomenclature hitherto used by me, I
called the motile filaments, microgametes; the pigmented cells, zygotes;
and the thread-like bodies, blasts. I also suggested a classification for the

* Owing to an error he thought that it was this kind in which I had first found the pig-
mented cells in 1897.

70

parasites of men and of birds. But there is still great divergence of opi-
nion on these subjects.

Evidently West Africa, a rich and enormous country hitherto para-
lysed by malaria, was destined to be the first objective. I lost no time
in urging the advisability of sending me there in order to complete my
studies of the disease and determine its agents on the spot. In July I
delivered my inagural lecture and demanded attention for my scheme for
extirpating malaria by attacking the pool-breeding mosquitoes [58]. At
the end of July 1899, accompanied by Mr. E. E. Austen of the British
Museum and Dr. H. E. Annet, Demonstrator of the Liverpool School of
Tropical Medicine, I left England for Freetown, Sierra Leone.

22. Sierra Leone; August— September^ i8gg. The Investigation Com-
pleted. — We have now reached the last chapter of this history — which
I fear has become tedious. If a literary simile may be allowed in a scien-
tific narrative, I had at least come to my Ithaca, after many mischances
sent by many opposing deities. Two years had elapsed since I had seen
the pigmented cells of the human parasites — two years of fruitless ef-
forts, interruptions and bad fortune; and seven years had elapsed since I
had commenced the special study of malaria; but now assisted by my
able colleagues and myself, I needed but a week or two to demonstrate
all the stages of the human parasites in dappled-winged mosquitoes, and
also to ascertain the fundamental principles upon which State sanitation
against tropical malaria should be based. I will be brief; the details are
given in the publications [60, 6j~\.

On the day after landing (loth August) we found two species of
dappled-winged mosquitoes (^AnopJieles costalis, Loew, and Anopheles fu-
nestus, Giles) in abundance. On the 13th August, we detected a pigmented
cell, evidently of the mild tertian parasite, in one of them. A few days
later, in some barracks where there was much malaria, we ascertained that
a quarter of the mosquitoes (almost exclusively A. cosialis) were infected;
and found in them pigmented cells evidently derived from all three varie-
ties of parasites — quartan, tertian and aistivo-autumnal. We also made
a few formal feeding experiments, and could have made as many more as
we pleased. The material was unlimited; but our time was short, and the
proof was already sufficient.

We then investigated the conditions under which the Anopheles breed
and propagate malaria. It was the rainy season and the place was full

71

of stagnant pools. Everywhere the larvae of the dappled-winged mosqui-
toes were in these pools, while those of the grey and brindled mosquitoes
occurred in tubs and pots. The great law of malaria — its connection
with stagnant water on the ground — was explained. Moreover, simply
by noting the presence of the larvae, we could tell at a glance which
pools were dangerous to health and should be dealt with in the public
interests.

The habits of the insects were noted and found to be precisely similar
to those of the Indian species. We studied particularly the characteristic
attitude of the larvae and adults of the dappled-winged mosquitoes, as
formerly observed in India — invaluable tests for the immediate and easy
recognition of the agents of the disease; we noted the evidence de-
monstrating the short flight of the insects, and their connection with rank
tropical vegetation; we disposed of the ideas that tidal swamps cause
malaria, but showed how earth-works produce outbreaks by the formation
of pools of rainwater. In fact we were able to give a thorough explana-
tion of the manner in which the old paludic and telluric theories of mal-
aria originated.

We were also able to establish for the first time the fundamental
principles which the State must adopt in order to extirpate malaria in
tropical cities. These are (i) scrupulous drainage of the soil; (2) pending
this, the persistent treatment of Anopheles breeding-pools by culicicides;
(3) the segregation of Europeans. We also recommended the protection of
public buildings, such as barracks, gaols, hospitals, and rest-houses by
wire gauze screens; the isolation of the sick; and the habitual employment
of mosquito-nets and punkahs by individuals.

Our results and recommendations were immediately communicated to
Government and also published in the medical press [60].

After our return to England in October we published a full report
of our experiences \p^'\. In this book, written by myself and endorsed
by my colleagues, I collected the principal results of all my researches
on malaria made during seven years; and illustrated the life-history both
of the human and avian parasites in mosquitoes by numerous photo-
micrographs made by myself. This work, therefore, which records the
completion of these labours by the successful demonstration of the whole
evolution of the human parasites in Anopheles, constitutes the summary
and conclusion of all my previous papers. It has been said that the book
was based on the writings of those who, as a matter of fact, learnt

72

everything from me; but I can say with exact truth that if no one except
MacCallum and Koch had touched the subject since 1895, scarcely a word
in the Report would have been different.

It should be added that in March 1900 I gave an abstract of the
history of my work in a lecture at the Royal Institution [68]; and par-
ticularly, that toward the end of the same year the president of the Royal
Society, Lord Lister, formally accepted my results in his address to the
Society. »

From this time my own effortts have been devoted almost entirely
to the practical campaign against malaria. Few people are aware of the
fact that even the most solid discoveries of science may be allowed by
the public to remain quite disused and inoperative unless strenuous efforts
are made to urge them upon the popular attention. Even yet, in spite
of the constant endeavours of many persons, very little has really been
done towards the extirpation of malaria. This has been principally due
to the fact that, for some inexplicable reason which wholly escapes me,
the chief prophylactic measure recommended by me, namely a campaign
against mosquitoes by drainage and petrolage, has been generally held
to be impossible; yet it is the only general prophylactic measure possible
in tropical towns. The struggle over this matter has been almost as
severe as that over the original problem ; but it is now drawing to a close.
It is impossible to discuss the matter here. Suffice it to say that in
the two principal towns, Havana and Ismailia, in which the measure has
been adequately employed, the reduction of malaria has already been as
much as eighty per cent.

This then is the conclusion of the history. I fear that some of the
personal details may have appeared out of place in the narrative; but
they have been introduced — though unwillingly — for a special reason.
No form of enterprise is of such transcendent importance to humanity in
general as the investigation of disease — the principal enemy of every
man. The interests of all nations, not only in the present but in the
future, demand that every possible encouragement should be given to such
investigations — particularly that medical men, who are in an excellent
position to undertake them, shall receive the warmest assistance in their
self-imposed task. The story, however, which I have felt it a duty to re-
cord in this lecture adds but one to the many instances of medical history
which show that little attention is given to this point. My labours will
be abundantly repaid if earnest students in this field of science receive, in

73

the future, in consequence of this narrative, a httle more assistance thau
was given to me.

23. Confirmation and Extensions. It is impossible for me to describe
here, even in detail, the vast amount of work which has been done in
many parts of the world on the mosquito theory of malaria since 1899;
but it is necessary just to touch upon some of the more immediate veri-
fications of my observations.

(i). Undoubtedly the first verification was due to Koch and his
assistants [63, 64]. Professor Koch was kind enough to communicate to
me at my request, in a letter dated the 15th May 1901, the origin and
progress of his researches on the mosquito theory of malaria. He says: —

s>The idea that mosquitoes may be the cause of malarial infection
occurred to me on my first visit to the tropics in British India in the
winter of 1883 — 84, and since then I have always spoken in this sense in
my lectures and to my assistants. I have not indeed myself published
anything about these views; but you will find a notice in R. PfeitTer's
work Beitrdge zur Protozoen-Forschimg, Berlin, 1892 (near the end).

»The fact that malaria, when it occurs epidemically, is often confined
almost entirely to the children, the adults remaining free and therefore
having become immune, I discovered first in villages in Java which lie in
the valley of Ambarawa. That was at the beginning of November 1899.
I reported on it on the 9th of December 1899, and my letter was pu-
blished in the Deut. Med. Woch. No. 5. 1900, beginning of February. I
obtained my first, successful cultivation of Proteosoma in mosquitoes in
company of Prof. R. Pfeififer in Rome in September 1898. We continued
the investigation in Berlin; and in the middle of November we followed
the developmental stages of the parasite up to the sickle-shaped bodies
in the poison glands of the mosquito — that is up to the end. We were
able to determine the form of iviirmchen (vermicule) in Proteosoma so
easily because I, with Professor Kossel, had already in June of the same
year (1898), without knowledge of MacCallum's investigation, detected the
origin of the spermatozoa, the process of fertilization, and the formation of
the iviirmchen in Halteridium.

^The publication of this investigation was very much delayed in con-
sequence of the long time taken for the reproduction of the photographs
in a way which satisfied me.

74

»At all events I have not thought it necessary to attempt to assert
my priority on this occasion as the matter concerned only the confirma-
tion of already known things. »

Professor Koch has the honour of having been one of the first, not
only independently to conceive the mosquito theory of malaria, but also
to attack it by experiment. He and Kossel independently observed the
function of the motile filaments by the employment of correct methods
of staining: this is practically admitted by Bignami (Lancet 1898, Vol II,
page 1898), who was later able, probably through his instruction, to de-
monstrate the chromatin in the motile filaments [50] — a thing which he
had refused to credit before. Koch also was the first to fill a gap in my
own researches on Proteosoina by demonstrating the passage of the ver-
micule through the wall of the mosquito's stomach — a subject in which
Grassi merely followed him later. That he succeeded in cultivating Proteo-
soma in Rome and Berlin, in September to November 1898, shows that
he was the first to confirm my own observations. About that time Dr.
Annett of the Liverpool School of Tropical Medicine saw some of his
preparations of pigmented cells in Berlin. Koch's discovery of the fre-
quent infection in native children in the tropics was one which I had en-
tirely failed to make — although I should have made it; and is an addition
to our knowledge of the very highest importance, being of far greater
intrinsic value than much of the trifling matter which has been put for-
ward in other quarters with much rHlaine. It enables us to explain with
ease the source of most malarial infections in the tropics, and, besides,
gives a complete revelation regarding the possibility of immunity in mal-
aria, a thing in which no one would previously believe. Added to this
Professor Koch has pressed still further onwards, and pushed with au-
thority and ability the great subject of the practical prevention of the
disease in the tropics — a matter the importance of which few writers
on the subject have seemed able to comprehend. The methods recom-
mended by me consist principally of the use of mosquito nets and the
extirpation of mosquitoes; but Koch at once inaugurated a new conception
which had not occurred to me and which consisted in the cinchonisation
of the people in malarious localities. Although this measure is not always
possible in its full extent, still experience shows that in a modified form
it is most useful; and I have come to the conclusion that it should al-
ways be enforced as much as practicable in addition to the measures
which I advocate. It was also Koch who was the first to call general

75

attention to the important fact that a sudden and ill-advised dose of qui-
nine is apt to precipitate attacks of blackwater fever in certain persons and
localities.*

(2). Many erroneous ideas have been formed about the Italian work (re-
ferred to in section 20) by those who have no practical knowledge of
malaria or full acquaintance with the literature. The facts are exactly
as follows.

The South of Italy affords unparalleled advantages for the study of
malaria, because abundance of material is there combined with great
facilities in connection with laboratories, literature, and scientific communion;
hence, though the principal discoveries have been made elsewhere, the
writers of Southern Italy have been able to add to them much detail,
which has proved more or less correct. It was hoped after Laveran's
discovery that they would be able to find the extracorporeal phase of the
parasite; but, unfortunately, misled by fondness for hypotheses, they fell
into fundamental errors. Most of them hastily concluded that the motile
filaments are »agony-forms»; and, as described in sections 6 and ii, A.
Bignami rejected Manson's induction on this account; and another writer,
G. B. Grassi, abandoned the whole mosquito-theory (i) because mos-
quitoes do not bite birds, (2) because they abound in places where there
is no malaria, and (3) because the malaria parasites die in the stomachs
of mosquitoes. At the same time he maintained that the extracorporeal
stage of the parasite is a free-living amoeba [10]. Bignami however, while
rejecting Manson's version of the theory, adopted King's, and stated that
he had even made some experiments on the subject in 1894; but these
were only referred to as a past event [29], and seemed to have been
quickly abandoned.

It is probable that these writers would have remained indefinitely in
this position but for the researches of others. In 1895 — 97' Sacharoff [23],
Simond [35], MacCallum [36], and myself [32] destroyed the Italian theory
regarding the motile filaments; and then the publications of the i8th De-
cember, 1897 [38], the 26th February 1898 [39], the 21st May [42], the
i8th June [41], the 24th September [43], and the nth October [46], com-
pletely demonstrated the life-history of this group of parasites in mos-
quitoes; clearly indicated the genus concerned in the propagation of
aestivo-autumnal fever; and gave other details mentioned in section 17.

* I should like to add, in contradiction of many inaccurate statements which have been
made, that his acknowledgment of my own observations has been the most complete possible.

7^

As all these papers, except those of the 21st May and the nth Octo-
ber (which were in fact covered by Hanson's papers of the i8th June and
the 24th September), were published in such a prominent organ as the
British Medical Journal, it is to be assumed that they were from the first
known to the Italian writers, who have always shown a prompt knowledge
of the labours of others. In his first publication [44], Grassi refers to my
work without mentioning my name or giving references — as if it were
then perfectly well-known in Italy. The later publications of Bignami and
Grassi [48, 47] show that they were quite intimate with it before they
themselves attained any definite results.

Such being the case, in order to follow my work in Italy and else-
where, all that was now needed w^as to determine the genera of my grey
and dappled-winged mosquitoes from such indications as I had been able
to give. The former had been described in two papers, and was most
evidently closely allied to Citlex pipiens\ and the latter, in which the
sestivo-autumnal parasites had been shown to develop, were described in
1897 [38] 3-S follows: —

»The latter are a large brown species biting well in the day-time, and
incidentally found to be capable of harbouring the filaria sanguinis hominis.
The back of the thorax and abdomen is a light fawn colour; the lower
surface of the same, and the terminal segments of the body a dark cho-
colate brown. The wings are light brown to white, and have four dark
spots on the anterior nervure. The haustellum and tarsi are brindled dark
and light brown. The eggs — at least when not properly developed —
are shaped curiously like ancient boats with raised stern and prow, and
have lines radiating from the concave border like banks of oars — so far
as I have seen, a unique shape for mosquito's eggs. The species appears
to belong to a family distinct from the ordinary brindled and grey in-
sects; but there is an allied species here, only more slender, whiter, and
much less voracious. » In the next paper [39] these small insects also are
called »dappled-winged».

At that time in Italy the CuHcidae had been carefully studied by
Ficalbi in several works [31], and it was an easy task for anyone pos-
sessing these works, and also having fresh mosquitoes for dissection, to
determine the genus of my dappled-winged mosquitoes from my description
alone. Although I did not give the entomological criterion of the genus
Anopheles (the long palpi of the female), I gave three other details which
sufficed for the identification. First, the dappled-winged mosquitoes be-

longed to a group distinct from the grey mosquitoes ((Tz//^,!- /z/z>;zjr type).
Secondly, both species of this group had spotted wings; and still more
particularly, one of them (certainly) had exactly »four dark spots on the
anterior nervure». Now it is well-known that very few Qilices and Stego-
myicE have spotted wings, while Anopheles almost alwa5^s have them.
The Anopheles, however, not only generally possess spotted wings, but the
spots are generally four in number and arranged along or close to the
anterior nervure. Lastly, if any doubt remained the observer would only
have to catch the first spotted-winged female gnat and to examine the
eggs within her, when they would be immediately seen to possess the
characteristic boat-like shape, with the well-known clasping membrane
simulating oars on either side.

It is curious that some of those who have written on the subject have
overlooked the fact that the very first Italian mosquito which from its
name alone w^ould be suggested by my description was Anopheles claviger.
Two of the synonyms of this insect are AnopJieles quadriniaciilatns, Say,
and Anopheles niaculipennis, Meigen!

There is, however, no doubt whatever that the Italians detected the
genus of my dappled-winged mosquitoes, because they themselves say so
in two of their articles of November, [48] and [51]. Nuttall admits the
fact [74]. But there is reason to suppose that they recognised the insects
long before November.

It was evidently Manson's paper of the i8th June 1898 which stimu-
lated the Italians to renewed activity, because they set to work shortly
afterwards. But their success was delayed by efforts towiirds originality.
Grassi endeavoured to find the guilty species of mosquito by its prevalence
in malarious localities. His efi"orts were a close repetition of mine in the
Sigur Ghat — even his servant was attacked by malaria as mine had been.
He discovered three species of guilty mosquito, namely Culex penicillaris,
C malaria; (so named by him — really C. vexajis), and Anopheles claviger
— principally (per lo meno) the first [44]. I have already shown in sec-
tions 1 1 and 1 2 how useless it is to attempt to identify the malaria-bearing
species by its preponderance in malarious places; and it has now been
demonstrated that even in Italy there is no such relation between the dis-
ease and its agent. Anopheles abound wdiere there is no malaria — even
round Liverpool. Needless to say then, two out of the three species
isolated by Grassi have nothing to do with the disease. He was right
regarding the third, A. claviger; but it is quite reasonable to suppose that

78

he detached this simply from my description of the dappled- winged mos-
quitoes. As a matter of fact all these epidemiological efforts of Grassi,
though interesting in a small way, were nothing but a series of vague
speculations.*

Meantime Bignami, after four year's inaction, had returned to his old
method of attempting to infect men by the bites of mosquitoes brought
from malarious places. His results are minutely recorded in his paper
[48]. He set to work in August — that is, after Manson had proclaimed
at the British Medical Association that I had succeeded in infecting birds
by the bites of mosquitoes [43]. Bignami's task was now vastly simpli-
fied; with the guidance of my work he collected his mosquitoes from in-
fected houses; whereas if he had continued to act in accordance with his
own theory he would have collected them from marshes — which would
have led to constant failure (section 13). He claimed his first success
early in November, but still could not say which of the various kinds of
mosquitoes employed by him had produced the result.**

Up to November therefore the Italians had failed either to find the
guilty species of mosquito or to demonstrate the life-cycle of the parasite
in the insects. At this point Charles's series of eight letters addressed from
Rome to me (dated from the 4th November to the 14th January) commence.
They have been printed by me with his consent; and show clearly
(what however can be also demonstrated from their own writings) that
the Italians were then intimately acquainted with my work; that they had
received my report [42] giving full details of technique: and that they had
detected the genus of my grey mosquitoes (from specimens sent by Man-
son) and of my dappled- winged mosquitoes (from my description). In his
letter of the 8th November 1898, Manson records having sent some of my
preparations to Charles and Bignami (on or before that date); and Charles
in his letter of the 25th November records showing one of these to Grassi
(on or before that date). It is possible, however, that the Italians had seen

* The writers of some zoological text books, who have evidently had little personal
experience of the disease, seem to have actually believed that Grassi determined the ^Ano-
pheles malariferi* by these efforts. That is not the case. In an early work [54] I said that
they were made independently of Manson and myself; but this was written before I studied
the Italian work with close attention; and since then I have withdrawn the statement [72].

** That human malaria is conveyed by the bites of mosquitoes had of course been
proved — practically to a certainty — by my infection of numerous birds three months previously.
Bignami's experiment was merely a formality of which the success could already be foretold
with confidence. The statement, frequently made, that he was the first to give experimental
demonstration of this fact may be set aside without comment.

79

my preparations long before this, as numbers of tiiem had been sent to
Manson and Laveran in the spring and summer; and they may also have
seen those of Koch, who had cultivated Proteosonia in Rome in September.

Bignami, Bastianelli, and Grassi had now evidently determined to re-
sort to the correct method for determining the guilty species of mosquito,
and imitated exactly the experiment by which I had ascertained the second
host of Proteosonia in the previous March. The experiment was recorded
by them on the 28th November [51]. They fed six Culex pipiens, one
Anopheles nigripes and four Anopheles claviger on some cases of crescents,
and at last found my pigmented cells in two of the last species. They
do not record the exact date on which this observation was made, but
from Charles's letters it would appear to have been on the 25th Novem-
ber or later.

This, if correct, was the first definite demonstration of the guilty
species of mosquito in Italy. It was made fifteen months after my original
demonstration of the same parasite in the same genus of mosquitoes in
Secunderabad on the 20th August 1897 [38], and nearly four months
after Manson had announced the whole life-cycle of Proteosonia at the
British Medical Association [43]. The Italian experiment was, however, of
doubtful correctness, because the authors do not state that the mosquitoes
used by them had been bred from the larvae [51]- At the same time they
actually impute to me the very fault which they themselves were com-
mitting, and do so contrary to the printed evidence of my own words.*

In their next paper [53] they claim to have found the various develop-
mental stages of the aestivo-autumnal parasites in A. claviger caught in
houses and stables, or fed on patients in hospital. Here again, examina-
tion of the publication shows that none of the insects employed seem to
have been bred from the larvae; and, what is still more important, the
number of insects on which the observations were made is not exactly
given. For all we know, the whole paper may have been written on the
strength of only a very few positive results; and this is the more po.ssible
because it describes a life-cycle which is an exact repetition of that of
Proteosonia. The authors give no precise differential experiments in order
to prove the connection between the pigmented cells seen by them and

* They say that my experiment was doubtful because my mosquitoes may have previously
bitten other animals [51, p. 314]. Now it is clearly stated in my publication [38, p. 1786]
that the insects used by me had been bred in »bottles from the larvae«; and from the whole
tenor of my researches it was evident that such was the case.

8o

the haematozoa. For this proof they rely upon my Proteosonia report [42],
to which, however, they scarcely refer. Although their paper gives to the
ignorant the impression of being original, it is in reality merely a re-
script of mine.

Meanwhile Bignami and Bastianelli had been continuing their attempts
to infect men by A. claviger taken from houses; and claimed a second
positive result early in December. In this case, however, by good fortune,
the infection proved to be a mild tertian one. Some months later, these
two authors published a paper [56] recording the development of this
parasite also in Anopheles. This was the first, and, mdeed, only important
Italian result which had not been previously indicated by me; I had made
no observation connecting the tertian parasite also with the dappled-winged
mosquitoes.

Subsequently the same authors and Grassi claimed to have demon-
strated the development of the parasites in Anopheles nigripes., A. bifur-
cattis^ and A. siiperpictiis.

They also claimed to have shown that the members of the old genus
Ctilex do not carry malaria; but this had long previously been abundantly
proved by me in India, at least with regard to the ^estivo-autumnal para-
site; and the fact is that Grassi had only identified my grey mosquitoes,
which I had shown to be negative to this parasite. Their first drawings
of the parasites in mosquitoes were not published until the spring.*

In my first reference to the Italian work I accepted it with some re-
serve; but after a careful examination of their writings made in 1900 I
felt much more scepticism. Their work during the winter of 1898 — 99
is evidently hasty and deficient in exact details regarding the various ob-
servations; and the general tenor of their historical passages is so inaccurate
as to inspire grave doubts regarding the whole literature. I think that at
that time they found my pigmented cells in a few, possibly a very few,
A. claviger and that Bignami and Bastianelli also showed that the tertian
parasite develops in the same insects; but beyond this it is impossible to
speak with confidence. Many of their details also are derived from me.

My work was completed in the autumn of 1899 at Sierra Leone
(section 22) and was published immediately [60]. Many of the details are
incorporated in Grassi's book published in June next year [69]. This

* For an independent account of all these researches the detailed history of Nuttall [65]
and especially his critique on the priority question [74], should be consulted.
See also [72, 73, 76].

work, which is dedicated to Manson, is principally a compilation of the
researches of others — the historical passages being quite inaccurate. At
the bottom of page 31 of the first edition, the author says, »Giova infine
far risaltare che io arrival agli Anopheles malariferi indipendentemente da
Ross, le cui ricerche sui parassiti malarici degli uccelli furono pubbHcate
quasi contemporaneamente alia mia prima Nota preliminare.» He and his
colleagues found the ■>^Anopheles malariferi» in Italy by detecting the genus
of my dappled-winged mosquitoes; but they did not incriminate it with
certainty until the end of November, five months after Manson published
my work on the malaria of birds [41]. Grassi's »first preliminary note»
[44] was published more than three months after this paper of Manson's,
and, moreover, refers to my work as a well-known matter even then, I
found the y> Anopheles malariferi» in two species of mosquito in \nd\2i fifteen
months before Bastianelli, Bignami, and Grassi found it in Italy. Speak-
ing quite strictly and accurately it is the principal merit of Grassi to
have discovered, not the y>Anopheles malariferi->, but its correct entomo-
logical name.*

Excepting the discovery of the host of the tertian and perhaps the
quartan parasites, the Italian work was simply a local aff"air, done, hke the
work of my colleagues and myself in Sierra Leone and of other observers
in many parts of the world, on the basis of my Indian researches culmi-
nating in July 1898 (section 17).

Of the sixteen and more species of Anopheles which have now been-
definitely connected with malaria, only three or four were incriminated by
the Italians; it is therefore quite incorrect to attribute the determination
of this relation to them — much more to attribute it, as some have done,
to Grassi alone. The connection between Anopheles and malaria has
been determined by the united eiiforts of many observers in many parts
of the world.

* Early in 1 903 this writer published a pamphlet purporting to be a translation of im-
portant papers on the subject {Documenii riguardanti la storia delta scoperta del modo di
transmissione delta malaria umana; Milano). It contains no bibliography nor accurate history
of the events; and omits most of the principal publications of my work. It purports to give
in full my paper recording the original discovery of the pigmented, cells [38]; but on examining
this copy I find that the drawings of the cells given by Manson, and the remarks of Manson,
Bland Sutton, and Thin (all of which, of course, absolutely establish the genuineness of the
discovery) are omitted without the smallest explanation. The author then proceeds to claim
the discovery for himself. This work also is dedicated to Manson — a fact which may lead
many to believe in its accuracy: but Manson has publicly stated that it was dedicated to him
without his permission (Lancet and British Medical Journal, 28th March, 1903).

Researcties on Malaria. 6

82

(3). Certainly not less important than the Italian work has been that
of the Malaria Commission of the Royal Society, consisting of Drs. Daniels,
Stephens, and Christophers. After confirming my results in Calcutta as
mentioned in section 20, Dr. Daniels proceeded to British Central Africa,
where he met Drs. Christophers and Stephens, who had proceeded there
after a month's stay in Italy in the autumn. These observers had great
trouble at first in obtaining suitable cases for experiment, but finally
succeeded in doing so. Daniels confirmed our results in Sierra Leone, and
added many useful and interesting details. Stephens and Christophers
afterwards followed us in Sierra Leone and elsewhere in West Africa, and
then proceeded to India.

The researches of all these gentlemen are given in the admirable re-
ports to the malaria committee of the Royal Society [71]. These researches
have had the effect of completely consolidating previous work on the
subject. The authors have shown no less than eight species of Anopheles
to be amenable to the malaria infection, and that Ciilices and StegoinyicB
are always refractory; they have demonstrated many of the habits of these
insects in various parts of the world; and, besides, have given us much
invaluable information regarding the pathology of the disease, especially of
blackwater fever. Stephens and Christophers also found independently the
great law of Koch regarding the prevalence of malaria amongst native
children in the tropics.

The perusal of the writings of these gentlemen and of many other observ-
ers will convince any one that it is impossible to do justice to them in the form
of a brief review; it is scarcely even fair to attempt to describe such la-
bourious work in a few words; and I shall therefore now draw this lecture to
a close with the remark, that I hope soon to deal with all these investiga-
tions in a manner which is due to them. But I should like to conclude
with the names of a few of those who have more recently added valuable
information to our store of knowledge regarding the mosquito theory of
malaria — particularly Ziemann, Manson, T. Manson, the members of the
numerous expeditions of the Liverpool School of Tropical Medicine (Drs.
Fielding-Ould, Annett, Dutton, Elliott, Logan Taylor), Fernside, James,
Low, Sambon, Van der Scheer, Van Berlekom, CelH, Nuttall, Shipley,
Ruge, Howard, Theobald, Schaudinn, and Sir William MacGregor. I omit
to refer to their works, and those of many others, only because it is im-
possible to do so properly within the limits of this work.

83

It would not be right, however, to conclude without referring to those
most conspicuous examples of the success of anti-malaria measures for
the improvement of public health — the cases of Havana and Ismailia.
A campaign against yellow fever and malaria was commenced at Havana
early in 1901 ; and Colonel and Assistant Surgeon General Gorgas of the
United States army, who was in charge of the work, has recently reported
as follows on the success of it, in a lecture delivered on May 22nd at
the New York Post Graduate Clinical Society.

»The results of these combined measures were very marked. Mos-
quitoes entirely disappeared from many parts of the city, and were decreased
everywhere. On the first inspection made in January, 1901, 26,000 col-
lections of fresh water were found in the city, containing mosquito larvae,
this exclusive of the cess-pools. In January 1902. the consolidated in-
spection reports covering the same area, showed less than 300; but the
most striking evidence was its results on yellow fever. It must be borne
in mind that yellow fever had been constantly in Havana since 1760, that
it was not, as it had been in our North American cities, some years
present and some years absent, but steadily every year and every month
and every day, in all that time. The deaths from yellow fever had been
since 1889, about as follows: — 1890, 303; 1891, 364; 1892, 352; 1893, 482;
1894, 388; 1895, 549; 1896, 1355; 1897, 743; 1898, 127; 1899, 118; 1900,
301; 1901, the first year of our mosquito work 5, and since September

1901, not a single case.

The work with regard to malaria is not quite so striking, and this
necessarily follows from the nature of the disease. But I think the results,
as shown by the sanitary reports, are very hopeful, with regard to malaria,
and indicate that in the course of time, malaria can be also eradicated.
In 1900, the year before mosquito work, the deaths from malaria were
344; in 1901, the first year of mosquito work, they had fallen to 151; in

1902, the second year of mosquito work, they had dropped to 90, and,
for the first four months of 1903, i6.»

At the end of 1902, the Suez Canal Company asked me to go to
Ismailia on the Suez Canal, in order to advise regarding the best mea-
sures to take against the malaria which had long been prevalent in that
town. I advised active operations against the mosquitoes; and this advice
was followed with great energy and success. The following table of
statistics, kindly suppHed by Prince D'Arenberg, the President of the Com-
pany, speaks for itself.

84

Statistiqiie du paludisme a Ismailia.

(From the statistics of the Suez Canal Company).

Mois 1S97 1898 1899 1900 1901 1902 1903 1904

Janvier 83 94 201 156 128 162 13

Fevrier 103 83 165 139 83 105 20

Mars 129 126 129 266 99 loi 16

Avril 135 127 109 175 100 64 14

Mai 173 77 126 169 82 133 9

Juin 180 43 126 114 68 154 15

Juillet . . . ... . . 188 81 104 145 74 120 23

Aout 242 86 107 166 123 130 19

Septembre 336 128 128 258 244 176 25

Octobre 254 178 172 228 372 139 39

Novembre 178 271 209 244 352 * 174 12

Decembre ..'.... 88 251 208 182 265 73 4

2,089 1,545 1.784 2,284 1,990 1,551 209

* Operations commenced.

Not only, however, from this place do we hear of reduction of sick-
ness and mortality. Undoubtedly the whole West Coast of Africa is much
improved, and good accounts continue to flow in from Lagos, the Gold
Coast, British Central Africa, Hongkong, and further India; and it is to
be hoped that within a few years malaria will, as Sir William MacGregor
says, have lost its terrors, at least for Europeans who are called upon to
serve in the tropics.

REFERENCES.

(This list of works, chronologically arranged, includes chiefly my own writings, many of which
are omitted in bibliographies, and such others as are referred to in the text. R. Ross.)

1. Manson. On the Development of Filaria Sanguinis Hominis and on the

Mosquito Considered as a Nurse. Linnean Society 1878. Also Trans-
actions Pathological Society 1881, XXXII.

2. King. Insects and Disease, Mosquitoes and Malaria. Popular Science

Monthly, New York, September 1883.

3. Laveran. Traite des Fievres Palustres. Paris 1884.

4. Celli & Marchiafava. Fortschritte der Medicin, 1885.

5. Macloskie. The Poison Apparatus of the Mosquito, American Natu-

ralist, 1888.

6. Lewis. Physiological & Pathological Researches. Published by the Lewis

Memorial Committee, London, 1888.

7. Agenore. Acqua Potabile e Malaria. Atti Delia Reale Accademia Medica

di Roma, Vol. V, Serie II, 1890.

8. Marino. Dell' Acqua dei Luoghi Malarici. Rif. Medica, 1890.

9. Grassi & Feletti. Several Papers. Centr. f. Bakteriologie, Bd. IX, 1891.

10. Grassi & Feletti. Contribuzione Alio Studio dei Parassiti Malarici. Atti.

dell' Accademia Gioenia di Scienze Naturali in Catania, Vol. V Serie 4 a.

11. Laveran. Du Paludisme et de son Hematozoaire. Paris 1891.

12. Ross. Fever with Intestinal Lesions. Transactions of the South Indian

Branch of the British Medical Association, 1892.

13. Ross. Cases of Febricula with Abdominal Tenderness. Indian Medical

Gazette, 1892, page 166.

14. Ross. Entero-Septic Fevers. Indian Medical Gazette, 1892, page 230.

15. Ross. A Study of Indian Fevers. Indian Medical Gazette, 1892, page 290.

16. Ross. Some Observations on Haematozoic Theories of Malaria. The Medi-

cal Reporter (Afterwards Indian Lancet), 1893, page 6^.

17. Ross. Nodulated and Vacuolated Corpuscles. The Indian Medical Re-'

cord, 1893, page 213.

18. Ross. Solution of Corpuscles Mistaken for Parasites. The Indian Medical

Record, 1893, page 310.

19. Smith & Kilborne. Investigations into the Nature, Causation and Pre-'

vention of Texas or Southern Cattle Fever. Bulletin No. i. Bureau
of Animal Industry, U. S. Dept. of Agriculture, Washington, 1893.
Also see Centralbt. f. Bakteriologie. 1S93.

20. Ross. Third Element of the Blood and the Malaria Parasite. Indian Me-

dical Gazette, January 1894, page 5. '

86

2 1. Ross. A List of Natural Appearances in the Blood which have been Mis-
taken for Forms of the Malaria Parasite. Indian Medical Gazette,
December 1894, page 441.

22. Manson. On the Nature and Significance of the Crescentic and Flagel-

lated Bodies in Malarial Blood. British Medical Journal, December
8th 1894.

23. Sacharoff. Ueber die Selbstandige Bewegung der Chromosomen bei Mal-

aria Parasiten. Centr. f. Bakteriologie, 1895.

24. Ross. Observations on the Crescent-Sphere Flagella Metamorphosis of the

Malarial Parasite within the Mosquito. South Indian Branch British
Medical Association, December 1895. Also Indian Lancet, 1896, pages
227 and 259.

25. Ross. Observations on Malaria Parasites made in Secunderabad, Deccan.

British Medical Journal, ist February, 1896.

26. Manson. The Life-History of the Malaria Germ Outside the Human Body.

British Medical Journal, 15th, 21st & 28th March 1896.

27. Ross. Some Practical Points Respecting the Malarial Parasite. Indian

Medical Gazette, 1896, page 42.

28. Ross. Dr. Manson's Mosquito Malaria Theory. Indian Medical Gazette,

1896, page 264.

29. BiGNAMi. Ipotesi dei Parassitti Malarici Fuori dell' Uomo. Policlinico, 15th

July 1896. Also English Translation Lancet, Vol. II 1896, pages
1363 and 1441.

30. Ross. Some Experiments in the Production of Malarial Fever by Means

of the Mosquito. South Indian Branch British Medical Association,
December 1896. (Read 30th October 1896). Also Indian Medical
Gazette.

31. FiCALBi. Revisione systematica d. fam. delle CuHcidae Europea, Florence 1896.

32. Ross. Observations on a Condition Necessary to the Transformation of

the Malaria Crescent. British Medical Journal, 30th January 1897.
2^. Ross. Further Observations on the Transformation of Crescents. South
Indian Branch of the British Medical Association, July 1897 (Rea-d 29th
January 1897. Also Indian Medical Gazette, January 1898.

34. Ross. Notes on Some Cases of Malaria, Amoeba coH and Cercomonas.

Indian Medical Gazette, May, 1897. (Proofs not corrected; full of
typographical errors).

35. SiMOND. L'evolution des Sporozoaires du Genre Coccidium, Annales de

I'lnstitut Pasteur, July 1897.

36. MacCallum. On the Flagellated Form of the Malaria Parasite. Lancet, 13th

November 1897. Also Journal E^xperimental Medicine, 1898, III.

37. Manson. A Method of Staining the Malarial Flagellate Organism. British

Medical Journal, 1897, II, p. 68.

38. Ross. On some peculiar Pigmented Cells found in Two Mosquitoes Fed

on Malarial Blood. British Medical Journal, i8th December 1897.

39. Ross. Pigmented Cells in Mosquitoes. British Medical Journal, 26th

February 1898.

40. Ross. Report on a Preliminary Investigation into Malaria in the Sigur

Ghat, Ootacamund. Transactions of the South Indian Branch of the
British Medical Association, February 1898. Also Indian Medical Ga-
zette, April 1898.
.41. Manson. Surgeon Major Ronald Ross's Recent Investigations on the Mos-
quito Malaria Theory. British Medical Journal, i8th June 1898.

87

42. Ross. Report on the Cultivation of Proteosoma, Labbe, in Grey Mosquitoes,

dated 21st May, 1898, Government Printing, Calcutta, 1898. Also
Indian Medical Gazette, November and December 1898. (In this copy
only one of the plates is given.) Second Edidon (Government Print-
ing), 1 90 1. (Many printers' errors).

43. Manson. The Mosquito and the Malaria Parasite. British Medical Journal,

24th September 1898. (Read at the Edinburgh Meering of the British
Medical Association at the end of July.)

44. Grassi. Rapporte tra la Malaria e Peculiare Insetti (Zanzaroni e Zanzare

Palustri). Dated 29th September, Pohcinico, ist October 1898.

45. Grassi. The same article as the above with the omission of certain pas-

sages, and undated. Atti della Reale Accademia dei Lincei — »per-
venute all' Accademia prima del 2 Ottobre i898».

46. Ross. Preliminary Report on the Infection of Birds with Proteosoma by

the Bites of Mosquitoes. Dated nth October, 1898. Government
Press, Calcutta.

47. Grassi. La Malaria Propagata per mezzo de Peculiari Insetti. xA.tti della

Reale Accademia dei Lincei, Seduta del 6 Novembre 1898.

48. BiGNAMi. Come si prendone le febri malariche. Ricerche Sperimentali,

Bulletino della R. Accademia Med. d. Roma. Dated 15th November
1898. Also translation in Lancet, 3rd and loth December 1898.

49. Grassi & DiONisr. II Ciclo Evolutivo degli Emosporidi. Atti della Reale

Accademia Dei Lincei. Seduta del 4 decembre 1898.

50. BiGNAMi & Bastianelli. On the Structure of the Semilunar and Falgellate

Bodies of Malarial Fevers. Lancet, 17th December 1898.

51. Bastianklli, Bignami & Grassi. Cultivazione Delia Semilune Malariche

dell' Uomo nell' Anopheles Claviger. Fabr. Atti della Reale Acca-
demia dei Lincei. Inviata il 28 Novembre 1898. Seduta del 4 de-
cembre 1898.

52. Grassi. Rapporte tra la Malaria e gli Artropodi. Atti della Reale Acca-

demia dei Lincei. Seduta del 4 decembre 1898.

53. Bastianelli, Bignami & Grassi. Ulteriori ricerche sul ciclo parassiti

malarici umani nel corpo del zanzarone. Atti della Reale Accademia
dei Lincei, Dated 22nd December 1898. Also later papers read on
the 5th February and the 7th May.

54. Ross. Du Role des Moustiques dans le Paludisme. Annales de I'lnstitut

Pasteur 1899, page 136. Presented to the Academic de Medecine,
24th January 1899.

55. Ross. Extermination of Malaria. Indian Medical Gazette, July 1899, 0^^'

port to Government of India dated the i6th February, 1899.)

56. Bastianelli & Bignami. Sullo Sviluppo dei Parassiti della Terzana. Bul-

letino della R. Accademia Medica di Roma, 1898 — 9, Ease. Ill dated
19th April. Also (with aggiunta), Annales d'Igiene Sperimentali 1899.

57. Grassi. Ancora Sulla Malaria. R. Accad. dei Lincei. Seduta del 18

giugno.

58. Ross. The Possibility of Extirpating Malaria from Certain Locahties by

a New Method. British Medical Journal, ist July, 1899.

59. Ross. Life-History of the Parasites of Malaria. Nature, 3rd August 1899.

60. Correspondent (R. Ross). The Malaria Expedition to Sierra Leone. Bri-

tish Medical Journal, 1899, Vol. II, 9th September, i6th September,
30th September, and 14th October, 1899.

61. Bastianelli & Bignami. Sulla Struttura dei parassitti malarici, e, in specie,

88

dei gamete dei parassite estivo-autunnale. Annales d'Igiene Sperimen-
tali, 1899.

62. Grasst, Bignami & Bastianelli. Ciclo Evolutive delle Semilune nell'

» Anopheles Claviger* ed altri Studi Sulla Malaria dall' Ottobre 1898
all Maggio 1899. Annales d'Igiene Sperimentali, 1899.

63. Koch. Ueber die Entwickelung der Malaria Parasiten. Zeitschr. f. Hygiene

und Infect. Bd. XXXII, 1899.

64. Koch. Berichte iiber die Thiitigkeit der Malaria Expeditionen. Deut. Med.

Woch. 1899 and 1900.

65. NuTTALL. On the Role of Insects, Arachnids and Myriapods, as Carriers

in the Spread of Bacterial and Parasitic Diseases of Man and Animals.
John Hopkins Hospital Reports, Vol. VIII. Also in Hygien. Rund-
schau, 1899.

66. NaTTALL. Die Mosquito Malaria Theorie, Centr. f. Bakteriol., 1899,

67. Ross, Annett, & Austen. Report of the Malaria Expedidon of the Li-

verpool School of Tropical Medicine and Medical Parasitology. Uni-
versity Press of Liverpool. Memoir II, February 1900.

68. Ross. Malaria & Mosquitoes. Nature, 29th March 1900. Also French

Translation, Revue Scientifique, 23 Juin 1900.

69. Grassi. Studi di Uno Zoologo Sulla Malaria. Reale Accademia dei Lincei,

4 giugno 1900.

70. Lord Lister. Presidential Address to the Royal Society on the 30th No-

vember 1900. -Extract in British Medical Journal, 8th December 1900.

71. Daniels, Stephens & Christophers. Reports to the Malaria Committee

of the Royal Society 1899 — 1903. Numerous papers.

72. Ross. Le Scoperte del Prof. Grassi Sulla Malaria. Two papers. Poli-

clinico, 1900 and 1901.

73. Calandruccio. Le Scoperte del Prof. G. B. Grassi Sulla Malaria, con

note ed aggiunte, 1900. Tip Barbagallo, Catania.

74. NuTTALL. On the Question of Priority with Regard to Certain Discoveries

upon the Etiology of Malarial Diseases. Quat. Journ. of Micros.
Science, 1901, page 429.

75. Theobald. A Monograph of the Cuhcidse or Mosquitoes. London 1901.

76. Ross. Die Entdeckungen des Herrn G. B. Grassi beziiglich der Malaria

und der Mosquitoes. Deut. Med. Woch. 27 Marz 1902 S. 231.

77. Giles. Report on Kala-Azar. Assam Secretarial Press, 1890.

78. Rogers. Report on Kala-Azar. Assam Secretarial Press, 1897. Also

See, Indian Medical Gazette, November 1897.

79. Ross. Report on Kala-Azar. Dated 30th January 1899, Government Press,

Calcutta i8qq.

DESCRIPTION OF PLATES.

(These Plates are copied from those given in my Report on the Cultivation of Pt'oteosoma, Labbe,

in Grey Mosquitoes, dated the 21st May 1898; but in the description I have substituted for

the appellation proteosoma-coccidia^ temporarily used by me for the pigmented cells, the word

now generally employed, namely zygotes. R. Ross).

Note. All the figures and plates were drawn by me accurately according
to scale from actual preparations, most of which were preserved in formalin.
Plates 11 to IX are faithful representations of entire fields.

Plate I. Drawings of zygotes from the second to the twelfth day.

Figures i — 5 zygotes of the second day. Figure 6, stained.
Figures 7 — 11 zygotes of the third day. Figure 12, stained.
Figures 13, 14, 15, zygotes of the fourth day. Figure 16, stained.
Figure 17, zygote of the fifth day.
Figures 18 — 22, zygotes of the sixth day and later.

Figures 23, drawing in outline of the stomach of a mosquito studded with
zygotes of the sixth day, seen by a low power.

Plates II — VIII. Fields of Leitz oel. imm. 170 minim. ''12th (inch).

Plate II. External coat of stomach studded with young zygotes of about 30
hours. Air vessel, crossing muscular fibres, and some oil globules, are seen.

Plate III. External coat of stomach studded with young zygotes of about 40 hours,

Plate IV. Zygotes of third day. Vacuolated forms.

Plate V. Zygotes of third day. Hyaline forms.

Plate VI. Zygotes of fourth day. One vacuolated and three hyaline forms.
One zygote of a second generation derived from a second feeding.

Plate VII. Zygotes of the fifth day.

Plate VIII. Zygotes of the sixth to seventh day. One zygote of a younger ge-
neration.

Plate IX. Pyloric end of stomach studded with zygotes of the seventh day,
seen by a power of medium strength.

Researches on Malaria.

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