56 
MALARIA 
blue. With the head and thorax pointing 
to the right, hold the mosquito in place with 
the needle in the left hand. Cut off the 
head with the curved needle held in the 
right hand. Press the thorax gently with 
the side of the curved needle, forcing a 
small mass of tissue from the neck opening. 
The tips of the glands may be seen pro¬ 
truding from this mass and are gently 
teased out. If they are not seen, tear the 
greenish-colored mass apart in search of the 
two glistening, blue-stained bodies. Trans¬ 
fer the glands, free of other tissues, by the 
curved needle tip to a clear drop of saline 
and cover with a cover slip. 
Examination of Fresh Preparations 
The preparations are examined with a 
compound microscope equipped with a me¬ 
chanical stage. 
Stomach. Focus a two-thirds objective 
on the stomach and move it to center of 
the field. Place the tip of a needle held in 
the left hand against the edge of the cover 
slip. Move the slide by means of the me¬ 
chanical stage against the needle point, so 
that the cover slip is held fixed. This causes 
the stomach to roll, so that all parts of its 
surface may be examined for projecting 
cysts. If suspicious bodies are found these 
are examined under a higher magnification. 
Small cysts are clear, round or oval bod¬ 
ies, without a retractile wall but with un¬ 
mistakable pigment granules. Large cysts 
show a distinct wall and only rarely is 
pigment visible. Mature cysts are finely 
striated, owing to the myriads of sporo¬ 
zoites which they contain. When only a 
few cysts are found they are usually in the 
posterior end of the stomach. 
The contraction of some of the circular 
muscle fibers in the central part of the 
stomach may produce externally extruding 
sacs of the stomach membrane that may 
cause confusion. Some of the fat cells con¬ 
taining fat globules might also be confused 
with cysts. Small cysts should never be 
diagnosed unless pigment is seen; large 
cysts are unmistakable. 
Salivary glands. Move the glands to cen¬ 
ter of the field of a two-thirds objective. 
With the tip of a needle press gently on the 
cover slip above so as to crush them. Then 
the fluid about the crushed glands is ex¬ 
amined under a one-sixth objective for 
sporozoites. These are slender, glistening 
rods, straight or slightly curved and about 
12 to 14 micra in length. Motility, at least 
in the sense of a translation of position, is 
questionable. 
Staining of Preparations 
Stomach. If a high incidence of infec¬ 
tion in the mosquitoes is expected, it is 
preferable to fix and stain the stomachs be¬ 
fore examination. Fixation and staining 
of specimens which have been for some time 
under a cover slip in saline is unsatisfactory. 
By staining freshly dissected stomachs in 
Mayer’s acid haemalum (formula follows) 
before examination, infections may be 
found which would be missed if examina¬ 
tion is limited to the fresh preparation. 
Cysts, while small, are detectable as early 
as the third day after an infective feeding 
in stained specimens, while such small 
cysts are very difficult to detect in un¬ 
stained preparations. 
After the stomach is removed and the 
Malpighian tubes trimmed off, it is covered 
with a large drop of Bouin’s fixative. 
Then cut off the fore gut and hind gut. 
Allow the fixative to act for 5 minutes. The 
specimen is transferred with a pipette 
throughout the different steps of the stain¬ 
ing process. After fixation the preparation 
is rinsed in distilled water to remove any 
excess of the fixative and is allowed to re¬ 
main in distilled water until all traces of 
the yellow color of the picric acid are gone. 
This requires several hours or preferably 
over night. From the distilled water the 
stomach is transferred to a 1:10 dilution 
of Mayer’s acid haemalum for an hour. 
The stomach is next washed to remove ex¬ 
cess of stain in a one per cent solution of 
acetic acid for 3 to 5 minutes, then trans¬ 
ferred to a one per cent solution of sodium 
carbonate to neutralize (the color changes 
to blue). For dehydration it is passed suc¬ 
cessively through 50, 70, 85, 95 and 99 per 
cent solutions of alcohol, remaining from 3 
