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10
Report by Dr. Andrewes on the Micro-organisms present in the Air op the
Central London Railway.
On March 8th, 1902, I was instructed, through the medical officer of the London County
Council, to conduct an investigation into the bacteria present in the air of the Central London
Railway.
My instructions were—To investigate twelve samples of air taken from different points in
the Central London Railway, and to compare them, bacteriologically, with twelve similar samples
taken at the same times from the fresh air outside; to enumerate the bacteria present, and, as
far as possible, to determine the species to which they belonged, employing, if need arose, suitable
means to test the pathogenic properties of any suspicious forms found. After consultation with
the medical officer and with the chemist to the Council, I commenced the work on March 14th.
The authorities of the Central London Railway were good enough to place every facility at my
disposal for carrying on the investigation. The large amount of time requisite for the growth of
the various cultures obtained, and for their determination, caused the work to be spread over some
four months, which, indeed, was of some advantage, as it enabled comparisons to be made under
varying seasonal conditions.
Method.—The method employed was, in principle, that introduced by Frankland, viz., the
aspiration of a known volume of air through a plug of glass wool, which retains all micro-organisms
and can subsequently be distributed through a suitable cultivating medium in a glass capsule.
This method has been used by Haldane, Laws and others with good results, and from my own
experience I can speak of it as simple and reliable. In my earlier experiments I used coarsely
pulverised glass wool alone, but I found, as others have found, that the addition of a small
quantity of very finely powdered cane sugar gave a more coherent and satisfactory plug, and one not
liable to charring during sterilisation if the temperature were not allowed to rise beyond 150° C.
I employed straight glass tubes about five inches in length, without any constriction. A suitable
amount of the powdered glass wool and sugar was introduced into these, and compressed by glass
rammers into a coherent plug about a quarter of an inch long. The coherence of each plug was
tested by forcibly aspirating air through it with a syringe. The ends of the tube were plugged
with cotton wool, one plug being introduced into one end, and two, an inner and an outer, into
the other. The tubes were then sterilised for an hour and a half in a dry-air steriliser at
13u°-150° C., and preserved in a bottle plugged with sterile wool.
For aspiration I used a large and well-made brass syringe fitted at the nozzle with a twoway
cock. The capacity of the syringe was 425 cubic centimetres, so that 11¾ syringefuls represented
very closely 5 litres of air. I graduated the stem of the piston, so that quarter syringefuls
could be measured accurately. I found this apparatus more convenient and portable than a
water aspirator, and I believe it to be equally accurate. The method of using it was as follows—
The tube containing the glass-wool plug was supported by two brass clips attached to rubber
suckers. The latter, when wetted, adhered firmly to any smooth wall or window. When no
suitable surface was at hand I attached them to a glass plate placed in a convenient situation.
The end of the tube provided with two cotton-wool plugs was attached by four feet of rubber
tubing to one of the nozzles of the syringe; this enabled me to stand at some distance from the
apparatus. The single cotton-wool plug at the free end of the tube was now removed with sterile
forceps and placed in a sterile capsule during the process of aspiration. A syringeful of air was
then sucked slowly through the tube. The air, entering its open end, passed first through the
glass-wool plug, next through the inner of the two cotton-wool plugs at the attached end, and then
through the outer cotton-wool plug. When the syringe was full, the tap of the two-way cock
was turned and the air expelled through the other nozzle. The tap was turned back, and a
second syringeful aspirated through the tube, and in this way 11¾ syringefuls of air were drawn
through the glass-wool plug. The cotton wool was now replaced in the free end of the tube; the
sample was duly labelled and returned to its bottle for transport to the laboratory. I found that
the process of aspirating 5 litres of air took about eight minutes.
The samples thus collected were treated in the laboratory in the following way—(1) The
outer of the two cotton-wool plugs in the end of the tube which had been attached to the syringe
was removed, and the inner plug was quickly transferred with sterile forceps to a culture tube of
broth. Had any micro-organisms failed to be arrested by the plug of glass wool and sugar they
must have been caught in this inner cotton-wool plug, which thus served as a test of the efficiencv
of the filtration. In the majority of cases the broth tube in which it was placed remained quite
sterile, indicating that all micro-organisms had been arrested. In those cases in which any
growth occurred in the broth, it occurred after several days, and was almost always limited to the
presence of a single species of bacterium or mould, indicating that only a single individual had
failed to be arrested. I have, therefore, every confidence that the cultures made from the glasswool
plugs fairly represented the real numbers of the organisms present in the 5 litres of air
aspirated. (2) A warmed sterile Petri capsule was now taken from the steriliser, and into it the
melted sterile culture medium (gelatin or agar-agar) was poured. The remaining cotton-wool
plug, i.e., the plug at the distal end, was removed from the sample tube, and the ends of the latter
passed through the flame of a Bunsen burner. With a sterilised glass rammer the plug of
pow dered glass wool and sugar was then forced out into the culture medium; it always came out
as a, solid cylindrical mass, which could easily be broken up with a sterile platinum wire and
evenly distributed through the melted culture medium. The sugar dissolved, and the tiny
particles of glass presented no obstacle to the counting of the bacterial colonies which subsequently
arose. When the culture medium had set firmly, the plates were put in the incubator—in the
case of the gelatin plates at 20° C., in the case of the agar-agar plates at 37° C. Here they were
incubated for four days, and the colonies were then counted. The period of four days was selected