A BACTERIOLOGIC STUDY OF OYSTERS, WITH 
SPECIAL REFERENCE TO THEM AS A 
SOURCE OF TYPHOID INFECTION. 
BY 
CHARLES J. FOOTE, 
DEMONSTRATOR OF BACTERIOLOGY IN THE MEDICAL DEPARTMENT OF 
YALE UNIVERSITY. 
FROM 
THE MEDICAL NEWS, 
March 23, 1895. Reprinted from The Medical News, March 23, 1895.] 
A BACTERIOLOGIC STUDY OF OYSTERS, WITH 
SPECIAL REFERENCE TO THEM AS A SOURCE 
OF TYPHOID INFECTION. 
By CHARLES J. FOOTE, 
DEMONSTRATOR OF BACTERIOLOGY IN THE-WffClCAL DEPARTMENT OF 
YALE UNIVERSITY. 
At the suggestion of Dr. C. A. Lindsley, Secretary of 
the Connecticut State Board of Health, I have made 
some bacteriologic experiments with oysters and with 
the water in which they grow, in order to determine how 
far the bacteriologic evidence supports the idea that 
typhoid fever may be communicated by oysters. 
The conditions favoring the communication of typhoid 
fever by oysters require that the Bacillus typlii abdomi- 
nalis should live a few hours at least in the water in which 
the oysters grow, and that, having been received between 
the shells of the oyster, it should live in the substance or 
juices of the oyster until such a time as this is eaten. In 
this locality this time would be from a few hours to four 
or five days, as it is the custom to “drink” the oys- 
ters in the river after bringing them in from the sound, 
for a few hours or days before opening them, in order 
to clean them, lessen their salty taste, and fatten them. 
The work of other investigators on the vitality of the 
B. typhi abdominalis in salt water is not quite ap- 
plicable to the instance under discussion, inasmuch as 
the temperature of the water is not given nor the amount 
of salt; and indeed, in some instances, the experiments 
were made with a sterile salt-solution. Freytag found 
that the B. typhi abdominalis would live in concentrated 2 
salt-solution five months.1 Giaxa detected it in unsteri- 
lized sea-water after nine days from the date of infec- 
tion, in sterilized water after twenty-five days.2 
My experiments were made to find how long the B. 
typhi abdominalis would live in water having the same 
percentage of salt, the same temperature, and the same 
bacteria as the water in which the oysters were planted. 
On December 20th a liter of water was taken from the 
river X, at low tide, in a sterilized flask directly over the 
oyster-beds. The flask was carried immediately to the 
laboratory, and ten c.c. of a bouillon-culture of the B. 
typhi abdominalis were emptied into it. The flask was 
then put in a wooden box outside of the window of the 
laboratory. Thus the temperature of the water ap- 
proximated at least that of the river. The mean daily 
temperature of the weather during the experiment was 
taken from the weather-reports, and was as follows: 
December 21st, 36° F.; 22d, 420 F.; 23d, 240 F.; 24th, 
240 F.; 25th, 38° F.; 26th, 240 F.; 27th, 250 F.; 28th, 
io° F.; 29th, 90 F.; 30th, 180 F.; 31st, 240 F. 
For the first few days the water was not frozen, pro- 
tected as it was in a wooden box, but during the last four 
days of the experiment there was a large cake of ice in 
the flask, and on December 30th the water was com- 
pletely frozen and the flask cracked. 
The last plate from the ice, made on December 31st, 
shows the vitality of the B. typhi abdominalis in frozen 
brackish water. 
Agar plates were made from the flask of water on 
December 23d, December 27th, and December 31st. 
The plates were kept at 3j° C. Check-plates were made 
also from a bottle of water from the river X, which was 
taken at the same time as the other sample, and was not 
infected with the B. typhi abdominalis. In every case 
1 Zeitschrift fur Hygiene, 1890. 
* Zeitschrift fur Hygiene, Bd. vi, p. 162, 3 
i c.c. of water was added to each plate, and in all 
cases in which I have stated that the B. typhi abdominalis 
was present cultures were made, from one or more 
colonies of each agar plate, on potato and in litmus 
milk, thus verifying the diagnosis. 
The results were as follows : 
Number of Colonies in c.c. of Water. 
Plate of 23d, of infected water, Innumerable cols, of P. 
typh. abd. 
Check-plate of non-infected water, 2 or 3 cols., none resemb- 
ling B. typh. 
Plate of 27th, of infected water, Innumerable cols, of B. 
typh. abd. 
Plate of 31st, of infected water, Innumerable cols, of B. 
typh. abd. 
An estimation of the chlorin in the infected water was 
made, and there was found to be o.i per cent, chlorin or 
0.15 per cent, salt.1 
On January 15th two more samples of water were 
taken from the river X in sterilized liter flasks.2 To one 
of the samples 10 c.c. of a bouillon-culture of the B. 
typhi abdominalis were added ; the other sample was 
used to make check-plates. 
The flasks were kept at the temperature of the labor- 
atory, as it was desirable to prolong the experiments, 
and it was feared that if exposed to the outdoor tem- 
perature the water would freeze and the flasks crack. 
The temperature of the laboratory during the experiment 
ranged from 6o° F. to 70° F. 
Plates were made at the dates recorded, with the fol- 
lowing results. 
1 For this analysis I am indebted to Prof. H. E. Smith, of the 
Yale Medical School. 
2 An analysis of this water showed it to contain 0.06 per cent, of 
salt. 4 
Water of river X—second sample—fa c.c. in each 
plate—water infected on January 15th. 
Water infected with 
Non-infected 
B. typh. abd. 
water. 
Plates of Jan. 16th, 
Innumerable cols, of B. 
4 cols ; none of B. 
typli. abd. 
typh. abd. 
Plates of Jan. 18th, 
Innumerable cols, of B. 
4 cols.; none of B. 
typh. abd. 
typh. abd. 
Plates of Jan. 21st, 
Innumerable cols, of B. 
5 cols.; none of B. 
typh. abd. 
typh. abd. 
Plates of Jan. 23d, 
Innumerable cols, of B. 
2 cols.; none of B. 
typh. abd. 
typh. abd. 
Plates of Jan. 28th, 
130 cols, of B. typh. 
3 cols.; none of B. 
abd. 
typh. abd. 
Number of Colonies in c.c. 
Plates of Feb. ist, About 10 cols, of B. Sterile. 
typh. abd.; many 
other kinds growing 
on plate. 
Plates of Feb. 5th, B. typh. abd. not dis- 
covered; plate over- 
grown with other 
kinds of bacteria. 
We may conclude from these experiments that even 
in extremely cold weather the B. typhi abdominalis will 
live in unsterilized salt or brackish water eight days at 
least, while in warmer water it rapidly diminishes in 
number after the first week, and cannot be detected in 
the water after three weeks. 
To determine whether the B. typhi abdominalis will 
live and multiply in oysters, it is necessary, as a prelimi- 
nary step, to find out what bacteria, if any, are found in 
oysters, and whether any of them resemble the B. typhi 
abdominalis. To determine this point some oysters 
were taken from the river X on November 27th, at low 
tide, scrubbed in city water, rinsed with sterilized water, 
dried on a clean towel, and opened with a sterilized 
knife. 5 
The juice surrounding the oyster was drawn off with a 
sterilized pipet, and gelatin plates were made, each 
one containing one drop of the juice. Tubes of Paretti’s 
solution were also infected with five drops each of oyster- 
juice. A sterilized platinum loop was plunged into the 
liver and stomach, twisted around, drawn out and rubbed 
over slant tubes of agar. 
The agar tubes and Paretti’s solution were incubated 
at 370 C., the gelatin plates at about 220 C. 
The oysters used in this experiment were numbered 
1, 2, 3, 4, and 5. 
The results were as follows : 
Tubes of Pare Hi's Solution. 
Oyster i Cloudy. 
• “ 2 “ 
“3 “ 
“4 “ 
“ 5 
Gelatin plates were made from each of the Paretti 
cultures, but no bacteria resembling in their growth on 
gelatin the B. typhi abdominalis were found. Most of 
the bacteria developing in Paretti’s solution from oyster- 
juice are anaerobic and are nearly all micrococci, so that 
they may be readily distinguished from the B. typhi ab- 
dominalis microscopically without making cultures on 
gelatin. 
The following is a brief description of the kinds of 
bacteria found in oyster-juice which grow in Paretti’s 
solution : 
(a) In gelatin tube-cultures this presents almost no 
surface-growth, but shows as a white streak along the 
line of puncture. There is no liquefaction, and the 
growth is very slow at 720 F. On potato it forms an in- 
visible growth. 
Grown in peptone-solution it gives no indol-reaction. 
It turns litmus milk slightly acid. It is a micrococcus, 6 
sometimes arranged in strings like a streptococcus. It 
is harmless when injected into the peritoneal cavity of 
white rats. 
(b) This bacterium grows dense, white, solid colonies 
on gelatin plates. The colonies are fairly round, but 
the outline somewhat dentated. After one week the 
colonies are nearly one-quarter of an inch in diameter. 
In gelatin tubes there is a thick, white, pasty surface- 
growth, with a good growth along the line of puncture. 
On potato it has a yellow growth. Under the micro- 
scope it shows large fat oval cocci. 
(c) On gelatin plates the colonies of this bacterium 
are small, round and white, of slow growth. There is 
no liquefaction. In gelatin tube-cultures there is no sur- 
face-growth, and only a few small yellow globules along 
the line of puncture. There is no change in litmus 
milk—no indol-reaction is produced. 
The microscope shows it to be a micrococcus. 
(d) In gelatin plates the colonies after one week are 
minute specks. Under a one-half inch objective they ap- 
pear as little, round, yellow drops. 
On potato at 370 C. there is a white growth after 
twenty-four hours. 
In gelatin tube-cultures there are little white specks 
along the line of puncture, with almost no superficial 
growth. After some days the gelatin becomes liquefied 
at the surface. 
The microscope shows medium-sized micrococci. 
Gelatin plates made with one drop of oyster-juice 
gave the following result: 
Oyster 1. Large number of small, white colonies. 
“ 2. Innumerable, mostly liquefying bacteria. 
“ 3. Innumerable. 
“ 4. A few colonies. 
“ 5. Innumerable. 
A few days later another experiment was made to 
determine more accurately the number of bacteria and 7 
the kinds found in oyster-juice. The gelatin plates 
were made with c.c. of juice in each plate. Nine 
oysters were examined. 
Number of 
Number of 
Number of 
bacteria in 
varieties 
liquefying 
Oysters. 
V20 c.c. of 
noticed 
in each 
Kinds recognized. 
juice. 
plate. 
I 
18 
5 
2 
2 
18 
5 
Many B. fluorescens 
liquefaciens. 
3 
12 
5 
B. gasoformans. 
4 
39 
3 
5 
43 
7 
5 
B. fluorescens 
liquefaciens. 
6 
48 
6 
4 
7 
12 
4 
2 
8 
84 
10 
11 
9 
34 
9 
At the same time agar plates were made with jjy c.c. 
juice of the same oysters, and the plates were incubated 
at 37° C. 
Agar Plates of Oyster-juice. 
Oyster 
I . 
Number of colonies in 
1/2o c.c. of juice 
growing at 370 C. 
I 
U 
2 . 
3 
“ 
3 • 
. . . . Sterile 
“ 
4 • 
. Many brown colonies 
“ 
S • 
8 
“ 
6 . 
. . . . . 2 
u 
7 * 
. . . . . 2 
8 . 
. . . . . 1 
it 
9 • 
. Many brown colonies 
To determine whether the contents of the stomach 
contained any bacteria, the oyster was cut open with a 
sterilized knife and the surface of the stomach scraped 8 
with a loop of platinum ; this was then drawn over a 
slant tube of agar. The loop of platinum was also 
plunged into the liver-substance, twisted around, drawn 
out, and rubbed over the surface of a slant tube of 
agar. 
Agar Tubes made by drawing a Loop of the Contents of 
the Stomach over the Surface of the Agar. Tubes 
incubated at 37° C., Bacteria in one loop growing at 
31° C. 
Oyster 1 Sterile. 
“ 2 . . . . . Sterile. 
“ 3 . . . . . Sterile. 
“ 4 2 cols., not B. typh. abd. 
“ 5 . . . . .1 col., B. megaterium. 
A second experiment resulted as follows: 
Oysters 
i 
Bacteria in loop of 
stomach-contents. 
. . I col. 
Bacteria in loop of 
liver-substance. 
Sterile 
2 
. . Sterile 
Sterile 
(< 
3 • 
. . Sterile 
Sterile 
(( 
4 • 
. . Sterile 
Sterile 
a 
5 • 
A few colonies A few colonies 
u 
6 . 
. . Sterile 
Sterile 
7 • 
. . Sterile 
Sterile 
<i 
8 . 
. . Sterile 
Sterile 
9 • 
A few colonies 
Sterile 
From these experiments we see that the number of 
bacteria in one cubic centimeter of oyster-juice which 
grow in gelatin range from 240 to 1680. The average 
number is about 684. 
It may be of some interest to compare the number in 
the oyster-juice with the number in the water taken 
directly over where the oysters were growing. Samples of 
water taken directly over the oyster-beds and secured at 
the same time that the oysters were taken out of the water 
contained on an average 9520 bacteria in a cubic centi- 
meter. Comparing this number with the number in the juice of the oyster we find that there are 8836 more 
bacteria in a cubic centimeter of the water surrounding 
the oyster than in the juice of the oyster. This indicates 
either that the juice of the oyster is destructive to certain 
bacteria or that the water undergoes a certain amount 
of filtration and purification in passing between the shells 
of the oyster. 
The kinds of bacteria growing at 22° C. from the 
juice of the oyster are in most instances the same as 
those found in the water in which the oysters grow. 
Thus we find, among the more common varieties present, 
the B. fluorescens liquefaciens and the B. gasoformans. 
In one instance the B. megaterium was isolated from 
the stomach of an oyster. This bacterium, as is well 
known, is found on cabbage-leaves. There is a kind 
of seaweed, called by oystermen sea-cabbage, which is 
abundant around oyster-beds and on oysters. Whether 
this explains the presence of the B. megaterium in oys- 
ters I do not know. 
Most of the bacteria found in the juice of the oyster 
do not grow at 370 C. By reference to the table of agar 
plates made from oyster-juice we see that there are 
rarely over 160 bacteria in one cubic centimeter of juice 
which grow at37° C., and often there are not more than 20. 
The examination of the stomach and liver of oysters 
shows in many cases that they are free from bacteria 
which grow at 370 C. The liver-substance was free in 
eight oysters out of nine, while the stomach was free 
from them in six out of nine. 
I have studied chiefly the bacteria in oysters which 
grow at 370 C., as most growing at a lower temperature 
may be ruled out as a cause of disease. The bacteria 
from oysters growing in Paretti’s solution I find, as a 
rule, quite different from those growing on agar plates. 
In the former case the bacteria are mostly anaerobic 
micrococci, while in the latter case they are aerobic 
bacilli. 
9 Some of those growing in Paretti’s solution, as already 
stated, resemble the B. typhi abdominalis by their in- 
visible growth on potato, but are easily distinguished 
microscopically. 
Two varieties isolated from agar plates liquefy gelatin 
rapidly, and are thus distinguished from the B. typhi ab- 
dominalis. 
Three non-liquefying kinds were isolated from the agar 
plates, which in the early stages of growth on gelatin 
somewhat resembled the B. typhi abdominalis, but were 
easily distinguished from it by their growth on potato 
and in litmus milk. 
Inasmuch as river-oysters, as a rule, are more exposed 
to contamination from sewage than oysters which are. 
taken four or five miles away from any such source of 
contamination, from deep water, it seemed of interest to 
compare their relative infectiousness. 
To do this bouillon-cultures were made by putting a 
half centimeter of the juice of an oyster into bouillon 
and allowing it to develop at a temperature of 370 C. for 
forty eight hours. One cubic centimeter of this culture 
was then injected into the peritoneal cavity of a white rat. 
Bouillon-cultures were thus made from seven river- 
oysters and from five oysters taken three miles out in 
Long Island Sound, south of the New Haven light- 
house. One cubic centimeter of each culture was then 
injected into the peritoneal cavity of white rats. After 
one month all of the five rats injected with bouillon- 
cultures from the sound-oysters were living and well; 
of the seven rats injected with bouillon-cultures of 
river-oysters five were living and well, but two had died. 
One died three days after inoculation, the other six 
days after inoculation. From the blood and the liver of 
one of the rats a bacterium was isolated showing a white, 
round colony on agar plates, in litmus milk producing 
an intensely acid reaction, and producing a yellow slimy 
growth on potato. From the blood, liver, and spleen of the other rat a 
bacterium was isolated, producing a brown growth on 
agar, with a green fluorescence. These experiments, as 
far as they go, seem to show a greater infectiousness 
of river-oysters. 
It is an easy matter to determine the fact that the B. 
tvphi abdominalis will liv; in oysters, but very difficult 
to tell whether it will propagate in them. A numerical 
estimation cannot be made of the number of bacteria 
in an oyster, even though we may know how many were 
introduced into it. It is clear, therefore, that conclusions 
in regard to number can be only very inexact. 
My method consisted in cleaning a number of oysters 
and then injecting one cubic centimeter of a bouillon- 
culture of the B. typhi abdominalis with a Koch syringe 
between the edges of the shells. In this way a pile of 
oysters were infected. The oysters were kept in a cool 
room, the temperature ranging from 50° to 65° F. The 
vitality of the oysters was not apparently impaired by 
the operation, for, when opened subsequently, most of 
them appeared fresh. 
Every few days one of the oysters was opened and 
agar plates were made from one half a cubic centimeter 
of juice. The surface of the oyster over the stomach 
was then scorched with a red-hot iron, and the stomach 
was opened with a sterilized knife. A loop of the stom- 
ach contents was then drawn over a slant tube of agar. 
The plates and tubes were kept at 370 C. 
In this way it was thought that if the B. typhi abdo- 
minalis multiplied, the agar plates and slant tubes for 
each succeeding oyster would show a greater number of 
the B. typhi abdominalis than the oysters opened before 
it; while if the B. abdominalis died, the juice of suc- 
ceeding oysters would show a quick decrease in number. 
Occasionally an oyster was opened which had lain in 
the same room with the other oysters, but had not been 
infected with the B. typhi abdominalis. Plates were 12 
made from the juice and slant tubes from the stomach, 
as a check on the method. 
As a preliminary experiment, some of the juice of a 
large oyster was drawn off with a sterile pipet, put 
into a sterilized test-tube, and infected with a loop of a 
culture of the B. typhi abdominalis. 
Every few days agar plates were made with two loops 
of this juice. The tube of juice was kept at a tempera- 
ture of between 6o° F. and 70° F. 
The results were as follows : 
Oyster opened and juice drawn off on December 21st. 
Number of colonies in 
Number of 
two loops of juice 
colonies in two 
after infection. 
loops of juice 
before infection 
Plate of Dec. 21st . 
. Innumerable colonies 
of B. typh. abd. 
Sterile. 
“ Dec. 23d . 
« i> •• <1 
“ Dec. 27th . 
a a it •< 
" Dec. 31st . 
. About 100 cols, of B. 
typh. abd. 
“ Jan. 10th . 
. One or two cols, of B. 
typh. abd. ; plate 
overgrown with 
other bacteria. 
The following is the record of results with living oys- 
ters : 
Oysters Injected with Bouillon-culture of B. typh. abd. 
on January 16th. 
Oyster A. Opened January 17th. 
c.c. juice contains innumerable cols, of B. typh. abd. 
Stomach, a few cols, of B. typh. abd. 
Oyster B. Opened January 19th. 
c.c. juice contains innumerable cols, of B. typh. abd. 
Stomach, 50 or 60 cols, of B. typh. abd. 
Oyster C. Opened January 22d. 
% c c. juice contains 200 cols, of B. typh. abd. 
Stomach, 2 or 3 cols, of B. typh. abd. 13 
Oyster D. Opened January 24th. 
% c.c. juice contains innumerable cols, of B. typh. abd. 
Stomach, about 14 cols, of B. typh. abd. 
Oyster E. Opened January 28th. 
A c.c. juice contains innumerable cols, of B. typh abd. 
Stomach, sterile. 
Oyster F. Opened January 28th. Check from pile not inocu- 
lated with B. typh. abd. 
A c.c. juice, sterile. 
Stomach, sterile. 
Oyster G. Opened February 2d. 
A c.c. juice contains about 60 colonies of B. typh. abd. 
Stomach, sterile. 
Oyster H. Opened February 13th. 
A c.c. juice contains about 40 cols, of B. typh. abd. 
Stomach, 3 cols, of B. typh. abd. 
In stating the number of colonies of the B. typhi ab- 
dominalis an exact number cannot be given, as there 
may have been in some instances other kinds of bacteria 
present which were mistaken for the B. typhi abdomi- 
nalis. In general, however, the number is approximately 
correct, as cultures were made from some of the col- 
onies of every plate, in litmus milk and on potato, and 
the mere plate appearance was not alone relied upon to 
determine whether I was dealing with the B. typhi ab- 
dominalis or some interloper. 
These experiments do not throw very much light on the 
question of the multiplication of the B. typhi abdomi- 
nalis in oysters. They do, however, seem to show that 
if multiplication does occur it takes place within the 
first two weeks, and that after that there is a progressive 
decrease in the number of the B. typhi abdominalis 
found in oysters, but that it may be found even after thirty 
days from the date of infection. They further show 
that the B. typhi abdominalis not only lives in the juice, 
but penetrates into the stomach and lives there for some 
time. In fact the B. typhi abdominalis lives longer in 
the juice and stomach of the oyster than it does in the 
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