in the same tube after use; but for clinical diagnostic purposes, they should be kept separate.
Preferable, the initial isolation of organisms
suspected of being Corynebacterium diphtheriae
should be carried out in a competent laboratory
near the source of the specimens.
To obtain a nasal and throat specimen, the swab
should be introduced into the nares at a right
angle to the plane of the face, inserted com-
pletely into the nasopharynx, and rotated over
the pharyngeal surface near or in the tonsillar
fossa. Since a nasal membrane is a particularly
good source of organisms, the swab should be
placed at the margin of the membrane, if present,
and the specimen taken without excessive bleeding.
To establish diagnosis in clinical diagnostic
work, the swabs should be gently rotated over the
surface of separate Loeffler blood serum slants
within two hours of collection, if possible, using
care not to break the surface of the medium. Long
delay in transporting such swabs to the laboratory
results in a considerable reduction of viable
organisms, and delays especially should be avoided
with swabs from convalescents and carriers. Primary
inoculation onto slants of a tellurite medium, if
it is to be used as a basis for morphological
diagnosis, may be misleading since tellurite has
a tendency to distort the morphology of the organisms
and the results can be confusing. However, plates
of cystine tellurite agar may be streaked with the
initial swab. In this way, a pure culture may be
obtained 2 to 4 hours earlier than plates inocu-
lated from the Loeffler slant following incubation.
In carrier surveys, both swabs may be streaked gen-
tly and repeatedly over the surface of a single
slant. Be sure to rotate the swabs between the
thumb and forefinger as the swabs are drawn back
and forth over the medium. Streaking of tellurite
plates with the initial swabs taken in carrier
surveys is of little advantage. In any case, if
the examination is not to be made locally, slants
only should be inoculated and forwarded to the
diagnostic laboratory together with information
as to how long they have been incubated.
On occasion, inhibitory lots of Loeffler's medium
have been encountered, due to the presence of
antibiotics in the serum from which the medium
was prepared. Therefore, precaution should be
taken to make sure that all lots of the medium
will support growth of C. diphtheria before they
are used. Diagnosis from smears made directly
from the patient should never be attempted
since many other organisms morphologically
resembling C. diphtheriae occur in the normal
or diseased throat.
In certain laboratories, procedures and media
varying from the above are routinely employed.
In such cases local preference should be deferred
to.
Bacteriologic culture is requisite to diagnosis of
gonorrhea in the female and desirable in gonorrhea
of the male. Recent and more dependable cultural
technics permit differentiating gonococcal from
non-gonococcal urethritis in the male and have
facilitated the diagnosis in the female. Diagnosis
by smear examination is simple but it is not as reliable
as culture. In chronic cases in both sexes and in the
detection of carriers, the culture method is far more
reliable.
The specimens submitted in acute gonococcal infection
for either film or culture are collected with a cotton-
tipped applicator from the urethra or cervix.
cultures are to be made, two sterile swabs should be
used and a tightly-stoppered tube containing 1 to 2
ml. of nutrient broth, or the transporation medium
of choice, is also necessary. The plating medium may
be inoculated directly from the patient. Otherwise,
place the swab in the tube of meat infusion broth in
which the gonococcus may be expected to survive 4 to
5 hours. This has the advantage of permitting delayed
plating, diluting the inoculum, and providing added
moisture to the surface of the solid medium.
In chronic
cases in the male, prostatic secretions and urine should
also be submitted.
In vulvovaginitis, specimens are obtained from the
vagina. Other sources of infectious material are
the conjunctiva, abscesses of Bartholin's glands,
the Fallopian tubes, pelvic lesions, and rectal
discharges. Cultures of blood, joint, and spinal
fluid occasionally reveal the gonococcus. Cultural
examination of the spinal fluid in cases of atypical
meningitis are desirable. In symptom-free patients
following treatment, isolations are often made from
the mucous membranes of the genitourinary tract,
from prostatic secretions, or from urine sediment.
Specimens taken as a test for cure must not be
taken less than 48 hours after termination of therapy
since residual sulfonamide or antibiotic inhibits
growth of the gonococcus.
If a competent local laboratory is available,
cultures should be carried out close to the
source of the specimen. But if culture is to
be delayed 18 to 24 hours after collection of
the specimen, a so-called "transportation"
medium is essential. For this purpose, semi-
solid gelatin blood agar and gelatin egg albumin
agar may be used.
(1) Cultures from the male
Aseptic technic is unnecessary but the speci-
men should be collected in as clean a manner
as possible. Pus or strippings from the penile
urethra are collected with the cotton swab; and
when no urethral exudate is present, the first
10 to 15 ml. of voided urine should be collected
since isolation from the sediment may be success-
ful.
In chronic cases, prostatic fluid should be
cultured directly onto a chocolate agar plate
or collected in a test tube containing 1 to 2
ml. of broth for later culture. If the quantity
is small, collect the exudate from the meatus
on a sterile swab.
(2) Cultures from the female
Sterile preparation of the vulva and douching
of the vagina are unnecessary. If the urethral
meatus appears normal and there is no exudate,
films and cultures from this source are not
indicated. In this instance, cultures are
made from the cervix only, using a bivalve
vaginal speculum, without lubricant other than
water or saline. After removal of the cervical
plug, compress the cervix with the blades of
the speculum to express secretions from the
endocervical glands. Collect the specimen
by inserting a small sterile swab into the
cervix 0.5 to 1 cm. In pregnancy, collect
the specimen from the cervical os. Culture
at once on chocolate agar or plate the swab
in broth or the transportation medium if culture
is to be delayed. For collection of specimens
from children and infants with vulvovaginitis,
a cotton-tipped applicator may be used; but
the use of a female glass catheter is preferable.
The film or culture is made from the vaginal
secretion which flows into the catheter.
Culture from miscellaneous sources
Aseptic technic is important in taking specimens from the anorectal area. After cleansing and application of an antiseptic, the specimens are taken from the terminal portion of the anal canal.
Pus from abscesses or the conjunctiva may be
inoculated directly or suspended in infusion
broth. Joint and spinal fluids should be
collected in sterile tubes without broth.
Blood for culture is obtained by venipuncture;
one ml. and 4 ml. amounts are added to 100 ml.
of glucose ascitic fluid broth; 5 ml. are added
to 2 to 4 ml. of 2.5% sodium citrate to prevent
clotting for use in blood agar plates.
Transportation of specimens. Urine, blood,
exudate in broth, spinal fluid, etc., should
be delivered to the laboratory and cultured as
soon as possible. No more than 6 hours should
elapse between collection and culture of the
specimen. If not cultured immediately from
the patient, the specimen should be refrigerated.
Under no circumstances should it be incubated
prior to cultivation.
5. Hemolytic Streptococcus Infections
The specimens usually submitted in suspected hemolytic streptococcus infections are material from the anterior nasal or nasopharyngeal areas, the throat, pus, sputum, spinal fluid, discharges, exudates, urine, blood, and milk. The technic of swabbing an area to be cultured is as important in the isolation of streptococci as is
the cultivation of the specimen taken. In strep-
tococcal infections of the upper respiratory tract,
the organisms may be cultured from the nose or
throat specimen. If obstruction is encountered,
force should not be used and the nasopharyngeal
specimen cannot be taken on that side. If only
one specimen can be taken, it should be a throat
culture. In carrier surveys, nasopharyngeal speci-
mens are preferable. Planting of swabs should be
accomplished within not more than 4 hours after the
specimen is taken. Milk specimens should be refriger-
ated from the time taken until they are cultured.
Initial isolation of streptococci should, if
possible, be carried out in a competent laboratory
close to the source of the specimen.
(1) Anterior nasal, nasopharyngeal, and throat
specimens. Material taken from the upper
respiratory passages is the most common type
of specimen cultured for hemolytic strep-
tococci. Individual sterile swabs are used
and anterior nasal specimens are obtained by
introducing the swab into the nares about an
inch. With the tip of the nose elevated, the
swab, moistened with broth or saline, is intro-
duced along the floor of the nasal cavity, under
the middle turbinate, to the pharyngeal wall.
Be sure to touch any exudate if present. With
the tongue depressed, pass a dry swab over the
tonsils and pharynx, being sure not to touch the
tongue. A method of transporting swab specimens
to the laboratory on filter paper strips may
also be used. The strips are inoculated with
the swab and air-dried. On arrival in the
laboratory, they are used to inoculate blood
agar plates by direct contact for several
hours. Spread the specimen over the surface
of a blood agar plate or Loeffler slant within
4 hours after it is taken, if possible; otherwise
place the swabs in tubes of broth for transport
to the laboratory.
(2) Pus, sputum, spinal fluid, discharges, exudates,
urine. A Gram-stain of the specimen at the
local or hospital laboratory will indicate
roughly the number of organisms present; and
if few, the fluid may be centrifuged and the
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