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Fluorescent antibody staining of organisms in nasopharyngeal
smears provides a rapid, specific diagnosis.
Leukocytosis (<100,000/ul) with marked
lymphocytosis (<90%), especially in patients over six months of age
Negative blood cultures
*Serologic tests are of little diagnostic
value because antibodies appear late. top
VIRAL PNEUMONIAS
LABORATORY FINDINGS
Isolation of influenza, parainfluenza, or respiratory
syncytial virus from nasopharyngeal or throat swab or from throat washings
Complement-fixing antibodies; these appear
8 days to 9 days after the onset of illness; antibody titre against
the specific causative virus shows a fourfold rise in serum specimens taken
two to three weeks apart
Fluorescent staining of viral antigens in nasopharyngeal
or bronchial epithelial cells establishes the diagnosis while the patient
is acutely ill
Sputum - no bacteria; polymorphonuclear, mononuclear,
and bronchial epithelial cells; the latter are those sloughed from
the infected air passages
White blood cell (WBC) count may be normal or decreased
with relative lymphocytosis. Leukocytosis above 15,000/ul
usually indicates the presence of a secondary bacterial infection.
Complement-fixing antibody - fourfold rise in titre
is diagnostic in serum drawn 3-4 weeks after onset. This is present
in 80%-90% of patients. If only convalescent serum is available,
a titre > 1:64 is suggestive of prior infection. Titres may remain
elevated for one or more years after infection. IgM antibody first
appears 7-9 days after infection, peaks at 4-6 weeks, and does not start
to decline until 4-6 months later.
Cold haemagglutinins - fourfold rise in titre during
illness is suggestive but not diagnostic. This is positive at the
seventh day, peak level is at four weeks, and it is negative by four months.
Only 25% - 50% of patients develop cold agglutinins; therefore, a
negative test does not exclude the disease. Many other conditions
show elevated cold agglutinin titres. These include infectious mononucleosis,
rubella, mumps, influenza, adenovirus, dysproteinemias, haemolytic anaemia,
paroxysmal haemoglobinuria, Raynarud's disease, and scleroderma.
Indirect fluorescent antibodies - rise in titre may
be demonstrated by this sensitive and specific test
Isolation of organism from sputum, throat washing,
or upper-respiratory-tract secretions
Sputum - leukocytes and monocytes; normal bacterial
flora on smear and culture
WBC - normal to 20,000/ul (average 10,000/ul
- 12,000/ul); predominant granulocytes.
Presence of characteristic organisms seen on carbol-fuchsin
or auramine-rhodamine-stained smears of suspected secretions or tissue
Characteristic cultures from involved sites of concentrates
of suspected material: sputum, bronchial washings, gastric fluid,
pleural fluid, urine, cerebrospinal fluid, pus, bone, marrow, endometrial
scrapings, tissue biopsy
WBC - usually normal, except for lymphocytosis or
monocytosis; leukemoid reaction or leukopenia may occur in miliary disease;
pancytopenia is suggestive of bone-marrow involvement
Moderate normocytic, normochromic anaemia in advanced
disease
Increased sedimentation rate in disseminated disease;
this is not an index of disease activity
Decreased serum albumin reflects severe chronic infection
Laboratory findings of extrapulmonary tuberculosis
(e.g. meningitis, renal disease, pleural or pericardial effusion, arthritis,
liver disease)
A definite diagnosis requires either repeated isolation
and identification of multiple colonies of the same strain of nontuberculous
mycobacteria or isolation under sterile conditions of the organism from
a closed lesion. These criteria are necessary to differentiate pathogenic
organisms from saprophytic organisms.
Actinomyces, Nocardia, and Streptomyces. These organisms have been shown to be higher
bacteria, but they were thought to be fungi for many years because they have
filamentous forms, 0.5 to 0.8 microns in diameter, which appear to branch. Some
species form aerial mycelia in culture. The clinical manifestations of infection
are similar to those of a systemic fungal infection. It is now clear that they
are not fungi but are closely related to the mycobacteria. Some facts that you
should know about these genera are that:
Actinomyces are anaerobic, while Nocardia
and Streptomyces are aerobic.
Nocardia stain partially acid-fast, Actinomyces
and Streptomyces are not acid-fast.
Actinomyces produce granules. Most
actinomycetes in tissue do not stain with the H & E stain commonly used for
general histopathology. All genera may produce granules; Actinomyces almost
always produce granules.
ACTINOMYCOSIS AND NOCARDIOSIS
LABORATORY FINDINGS
Actinomycosis
Organism is seen on Gram's stain or material from
infected sinus tracts, abscess cavities, or empyema fluid. The organism
may be found normally in sputum and the tonsils and pharynx.
Presence of yellowish particles (suffer granules)
within pus or infected sputum; the granules comprise Actinomyces
colonies
embedded in a matrix of calcium phosphate
Growth of organism on culture of pus or biopsy tissue
Organism shows gram-positive filaments and is weakly
acid-fast.
Growth of organism on Sabouraud's medium without
antibiotics and on blood agar; it usually grows very slowly, is often
mixed with other organisms and some strains may be killed by mycobacterial
concentration techniques; occasional strains may grow rapidly on
blood agar
Link to image source The yellow lesion is a severe histoplasmosis scar that involves the macula
The fungal organism has a propensity to result in ocular disease, however, and the macula is the most common site of infection. When macular infection occurs, the patient may experience visual loss due to inflammation and scarring of the macula, and some patients will develop hemorrhage in the macula, due to the growth of vessels beneath the retina (choroidal neovascular membrane). The presence of the macular lesion, and peripheral retinal lesions known to ophthalmologists as "Histo spots," is generally all that is necessary to make the diagnosis. There is no single test that proves the diagnosis, however, and in some cases, blood and skin testing, and a chest x-ray may help confirm the diagnosis.
LABORATORY FINDINGS
Culture of H. capsulatum from skin or mucosal
ulcers, sputum, blood, spinal fluid, bone marrow, or tissues is required
for definitive diagnosis.
Tissue biopsy of bone marrow, mucosal ulcers, liver,
lymph nodes, lung with identification of H. capsulatum
Polyvalent fluorescent antibody reagent for the detection
and identification of H. capsulatum
Complement fixing antibody titres - positive in 80%
of chronic cases, but in only 50% of acute cases; appear in 3rd to 6th
week; may persist for months or years if the disease remains active.
When paired acute and convalescent sera are available, a fourfold or greater
rise in titre is highly suggestive of recent infection. Titre above
1:32 is strong presumptive evidence of active infection. Because
of cross-reactivity, titres may be positive in blastomycosis, aspergillosis,
and coccidiodomycosis. The test thus has low specificity. False-positive
and false-negative results may occur with chronic cavitary and disseminated
disease. This test is more sensitive than the immunodiffusion test
in patients with subclinical infection.
Latex agglutination test - this is often positive
in acute disease because it detects IgM antibody. It is not dependable
in chronic infections because it becomes negative in 5 to 8 months, even
with persistent active disease.
Immunodiffusion and counterimmunoelectrophoresis
tests are useful screening procedures. Precipitin bands of diagnostic
value are designated h and m. The m band has
been considered presumptive evidence of infection with H. capsulatum.
If
the patient has not had a recent histoplasmin skin test, detection of an
m
band
may serve as an indicator of early disease, because this band appears before
the h band and disappears more slowly.
It is important to note that levels of complement-fixing,
precipitating, and agglutinating antibodies to H. capsulatum may
be significantly increased in histoplasmin-sensitized persons after a single
histoplasmin skin test.
Etiology
Coccidioidomycosis is caused by Coccidioides immitis, a dimorphic fungus that grows as a mold in the soil. The mold forms arthroconidia within the hypha, a type of conidia formation known as enteroarthric development (Figure 1) (4). C. immitis is the only species within the primary pathogenic fungi that has this type of conidia development. Alternate conidia undergo autolysis, leaving empty spaces between viable arthroconidia. The arthroconidia are released into the atmosphere when the wind ruptures the hypha. C. immitis infects humans and animals almost exclusively by the respiratory route (5). Once inhaled, the arthroconidia cluster in the lungs and undergo a dramatic morphologic change. The round cells, which develop into spherules, undergo repeated internal divisions until they are filled with hundreds to thousands of offspring, termed endospores. This process occurs over 48 to 72 hours (6). When the spherule ruptures, each released endospore has the capacity to develop into a mature spherule
LABORATORY FINDINGS
Identification of organism by direct or phase-contrast
microscopy of tissue or body secretions
Isolation of organism by culture of sputum, gastric
contents, cerebrospinal fluid, exudate, synovial fluid, tissue
Precipitin antibodies are IgM, appear early, and
are detectable by the 3rd week of illness in approximately 90% of symptomatic
patients; they are uncommon after the 5th month. The latex
agglutination test also detects IgM antibodies of early primary infection.
This test is more sensitive than the precipitin test but is less specific.
Neither of these tests provides information about disease severity or prognosis.
Complement-fixing antibodies are IgG, and they appear
later and persist long than do precipitin antibodies. They appear
4 to 5 weeks after exposure and decrease after 4 to 8 months, but they
may remain positive for years. The antibody titre correlates with
disease severity and response to therapy. These antibodies are positive
in approximately 50% of nondisseminated primary infections and in 96% to
100% of disseminated infections. Antibodies in spinal fluid indicate
involvement of meninges.
The immunodiffusion test, with antibody to the complement-fixing
antigen, is negative in early primary infection but is a good screening
test for other stages.
Each yeast is multinucleate with 2-5 nuclei per cell. The mechanism for the phase transition is unknown, but the reason for the transition is quite clear, as described below. The fungus can also form a sexual state; the so-called teleomorph form is Ajellomyces dermatitidis, which forms a gymnothecium. The gymnothecium is kind of like a cleistothecium (closed ascocarp) as found in powdery mildews or Aspergillus. The gymnothecium, however, has an outer covering that is more loosely woven, so that ascospores can fall out without having the covering degraded by other microbes.
-----------------------------------------------
Blastomycosis starts out as a lung infection caused by inhalation of the conidia. The conidia transform into the yeast form, The transformation into a yeast involves many physiological and biochemical changes, having the effect of "buying some time" for the fungus to become established. The fungus evades the body's immune system by changing its surface antigens. Once established, the infection progresses as an occult, insidious (hidden and sneaky) process or as a chronic, expanding, and eventually systemic infection. The patient may have severe presenting symptoms or may be inapparent infection that resolves spontaneously or disseminates to another location. If symptoms do appear, the incubation period may be long - 21 to 106 days. It starts with mild progressive respiratory infection with a dry cough, some pleuritic pain, hoarseness, and a low grade fever, symptoms that overlap many other diseases, such as tuberculosis or chronic histoplasmosis. If the primary pulmonary infection does not resolve, severe progressive blastomycosis can result. It can sometimes be fatal. Sometimes the disease spreads to the skin (cutaneous blastomycosis, as shown to the right), the bones (osseous blastomycosis), the urogenital tract, or the central nervous system. The tissue response is variable, but organism shows up as big blue broad based budding yeast. An unequivocal diagnosis cannot be made unless this form is seen. A diagnosis may also involve isolating the fungus and growing it at room temperature and body temperature.
LABORATORY FINDINGS
Identification of organism by direct or phase-contrast
microscopy of sputum, pus, or tissue
Isolation of organism by culture of sputum, pus,
or tissue is required for definitive diagnosis.
Complement-fixing antibodies may be positive or negative
with systemic infection. Cross-reaction with H. capsulatum further
detracts from the diagnostic usefulness of this test.
The immunodiffusion test for blastomycosis is specific
and has a sensitivity of approximately 80% Negative tests, therefore, do
not exclude the diagnosis.
Leukocytosis and increased sedimentation rate occur
in active disease.
Latex slide agglutination on serum and spinal fluid
specifically detects cryptococcal antigen. This is a screening test.
The titre is usually proportional to the extent of infection. This
is more sensitive in diagnosing cryptococcal meningitis than is the India
ink test.
Tests for cryptococcal antibody - indirect fluorescent
antibody test and tube agglutination test; the latter is more specific:
a positive antibody test may occur early in the course of the disease and
in localized infections: the antibody test may have prognostic value;
these tests usually become positive after the latex agglutination test
becomes negative.
Identification of organism on smear; culture
of organism from sputum or lung tissue; because the organisms are
usually saprophytes their isolation is not diagnostic but must be correlated
with clinical information. Diagnosis requires demonstrating the fungus
in
tissues.
Immunodiffusion test for antibody, when used with
reference sera, is 100% specific. Conversion of the immunodiffusion
antibody test from negative to positive is diagnostic of infection.
Demonstration of one or more precipitating antibodies indicates infection,
fungus ball formation, or allergy due to an Aspergillus species.
Laboratory findings of an underlying disease, such
as tuberculosis, asthma, bronchiectasis, or bronchogenic carcinoma
Sputum - usually yellow or white and mucoid;
it may contain eosinophils, crystals, and mucous casts; neutrophils and
bacteria indicate superimposed infection
Increased blood and nasal eosinophils
Early decrease in arterial PO2; there is a
greater decrease as the condition becomes more severe
Early decease in PCO2 as a result of compensatory
hyperventilation
PCO2 increases toward normal as compensation fails.
pH - increases early (respiratory alkalosis);
decreases as condition worsens (respiratory acidosis)
Theophylline blood levels are useful in monitoring
therapy.
Increased serum angiotensin-converting enzyme - this
enzyme converts angiotensin I to angiotensin II and normally occurs within
the pulmonary capillary endothelial cells. Increased activity of
this enzyme in sarcoidosis is believed to be the result of an increased
rate of enzyme synthesis by epithelioid cells of the carcoid granulomas
and of release of the enzyme into the bloodstream. In a recent report,
83% of patients with active disease had elevated enzyme levels. 10 The
enzyme level tends to fall during spontaneous or steroid-induced remission
of the disease.
Increased sedimentation rate and serum fibrinogen indicate active disease.
Decreased serum albumin reflects chronic disease.
Increased serum globulins; stepwise increase of Alpha2-,
Beta-, and y-globulins, especially IgG; this reflects
humoral immunologic response of chronic inflammation
Leukopenia (<5,000/ul) in 40% of patients
Mild normocytic anaemia - chronic-disease type
Decreased lymphocytes
Increased serum uric acid
Increased urine calcium as a result of increased intestinal absorption
due to increased sensitivity to vitamin D.
Laboratory findings reflecting specific organ involvement of lungs, liver,
spleen, central nervous system, pituitary, or kidneys
Effusions are often subdivided into transudates,
which are ultrafiltrates of plasma and have a low concentration of protein,
and exudates, which result from capillary damage or lymphatic blockage
and have a high concentration of protein, and exudates, which result from
capillary damage or lymphatic blockage and have a high concentration of
protein. The traditional differentiating factor between the two types
of effusions has been that an exudate has greater than 3.0 g/dl of protein
and a transudate has less than 3.0 g/dl of protein. This has resulted
in too great an overlap between the two categories. The following
criteria are now widely used to define an exudate:
1. A pleural fluid-serum ratio
of total protein of 0.5 or greater
2. A pleural fluid lactate
dehydrogenase (LDH) of 20 IU or greater
3. A pleural fluid-serum ratio
of LDH of 0.6 or greater
Values below these indicate that the fluid is a transudate.
Conditions causing pleural transudates are congestive
heart failure, cirrhosis, and hypoalbuminemia. The most common causes
of pleural exudates are pulmonary embolism and infarction, malignant tumor
involving the pleura, empyema, pneumonia, and tuberculosis.
Sputum - abundant, foul, purulent; Gram's stain and
acid-fast stain rarely show pathogenic organisms: cultures may show
one or more aerobic and anaerobic bacteria. Mycobacterium tuberculosis,
or
fungi
Blood culture may be positive in the acute stage.
Leukocytosis (15,000/ul) - 30,000/ul)
- this is often absent in elderly or debilitated patients.
Early decrease in arterial P02, which becomes more
marked as pulmonary oedema progresses
Arterial PC02 may be decreased, normal, or increased.
A decrease reflects compensatory hyperventilation and an increase reflects
impaired gas exchange in severe oedema.
Arterial pH is usually normal or increased,
but occasionally decreased. Increased pH usually reflects
decreased PC02; decreased pH reflects increased PC02 or lactic acidosis.
Decreased arterial P02 (usually to 60 mm Hg - 80
mm Hg) is the most constant laboratory finding
Decreased PC02 and increased pH (respiratory
alkalosis) due to increased ventilation as compensation for hypoxemia;
this depends on the size and duration of the infarction
Leukocytosis of up to 15,000/ul
Increased sedimentation rate
Increased serum LDH (especially LDH2 and LDH3) -
rises on the first day, peaks on the second day, normal by the tenth day
Determination of lecithin and sphingomyelin in
amniotic fluid is the single most accurate test of foetal lung maturity.
The lecithin - sphingomyelin ratio (L/S ratio) is lower in prematurity
than at maturity. RDS is unlikely when the ratio is high, indicating
lung maturity. When a mixture of amniotic fluid and ethanol is shaken,
the mixture will produce foam if an adequate amount of surfactant is present.
If amniotic fluid is not available, the L/S ratio may be determined through
analysis of the gastric aspirate of the infant.
RDS and ARDS
The following are cumulative findings occurring
in order of increasing severity of pulmonary insufficiency:
Decresed PC02 and oxygen saturation, probably due
to decreased ventilation-perfusion ratio and to shunts
Increased PC02 and decreased pH (respiratory
acidosis), due to decreased ventilation and impaired compensatory response
to hypoxemia
Decreased bicarbonate and greater decrease in pH
(metabolic acidosis) secondary to tissue hypoxia and lactic acidosis
Arterial P02 and PC02 are the only clinical laboratory
indicators of pulmonary function. (pH is altered with changes
in PC02, but this reflects changes in bicarbonate.) Decreased arterial
P02 is an early and sensitive indicator of respiratory insufficiency.
Hypoxema occurs in the following circumstances:
Hypoventilation: impaired movement
of air in and out of the lungs. This may be seen following sedative
or narcotic drug depression of respiration; impaired cerebral control
of respiration due to stroke, brain tumour, or injury; weakness of
respiratory muscles; or obesity.
Diffusion abnormality: impaired exchange
of 02 and C02 between alveoli and pulmonary capillaries. This may
occur owing to injury to the alveolar membranes, such as following prolonged
use of certain cytotoxic drugs.
Ventilation - perfusion abnormality: diminished
ventilation to areas of the lung that continue to receive normal blood
flow. Associated with this abnormality are many forms of lung disease,
including pulmonary oedema, viral pneumonia, pulmonary embolism, and RDS.
Right-to-left shunt: flow of venous blood
through portions of the lungs that are completely unventilated. This
could occur with lung collapse (atelectasis); extensive filling of
the lungs with oedema fluid, pus, blood, or inflammatory cells as in pneumonia;
or with accumulation of air or fluid in the pleural cavity.
The four abnormalities that cause hypoxemia may be
differentiated by determining the patient's arterial PC02 and P02 while
he is breathing room air and again while he is breathing room air and again
while he is breathing 100% oxygen. The basis for this differentiation
is shown in Table 2-1.
Arterial PC02 is the single most important measurement
of alveolar ventilation. C02 retention (hypercapnia) occurs less
frequently than does hypoxemia and indicates a serious ventilatory problem.
Hypercapnia is accompanied by acidosis and hypoxemia unless the patient
is receiving oxygen. This occurs acutely following sudden airway
obstruction, pulmonary oedema, sedative overdose, or cardiac arrest.
This also may occur following long-standing severe pulmonary or alveolar
hypoventilation, as seen in emphysema and chronic bronchitis.
Decreased PC02 (hypocapnia) is due to hyperventilation
and occurs more frequently than does hypercapnia. Hypocapnia is accompanied
by alkalosis. Hyperventilation is seen as a response to hypoxemia
in many pulmonary diseases including pneumonia, atelectasis, pulmonary
embolism, and acute RDS.
Table 2-1. Pulmonary Causes of Hypoxemia
and Their Differentiation
ABNORMALITY
ARTERIAL PC02
ARTERIAL P02 (ROOM AIR)
ARTERIAL P02 (100% 02)
Hypoventilation
Increased
Decreased
Normal
Diffusion abnormality
Normal or decreased*
Normal at rest: decreased during exercise
Normal
Ventilation-perfusion abnormality
Normal, increased, or decreased
Decreased
Normal
Right-to-left shunt
Normal or decreased*
Decreased
Low
Fall in cardiac output in patient with right-to-left shunt
Normal, increased, or decreased
Decreased
Low
*Attributable to hyperventilation from secondary causes
(Adapted from Hyde RW: Clinical interpretation
of arterial oxygen measurements. Med. Clin. North Am 54:617-629,
1970)
Nocardia
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