Understanding
Investigations III Microbiology
Different
methods are available to detect infection. Some are non-specific, such as
changes in blood inflammatory markers, and others give specific information
about the exact infection. Proof of infection includes direct detection (e.g.
microscopy, antigen detection, PCR), detection of an antibody respons ogy) or
culture of an organism froma normally sterile site.
Blood culture
Culture of a with of a bacterium (or occasionally a
fungus) from blood is usually definitive proof of infection. However, if the
culture is of several different organisms, or if low pathogenic bacteria
normally found on the skin (such as coagulase-negative Staphylococci)
are isolated, then the result should be interpreted with caution. The way the
blood sample is taken is crucial—the skin must be thoroughly cleaned with
antiseptic and an aseptic technique used. Very small blood samples (<1
mL) reduce the chances of a positive culture. Blood cultures usually take 24–
48 hours to show evidence of infection and so are usually used to confirm a
clinical suspicion of infection retrospectively. Cultures will provide information
about the sensitivity of the organism, which can be used to rationalize
antibiotic therapy. Most modern blood culture systems are able to detect
significant bacteraemia in children with a single medium, which is designed
specifically for this purpose. Unusual results should always be discussed with
a microbiologist.
Serological evidence of infection
Measurement of antibody response to specific infectious agents can be
useful. This is important to check for prior immunity (e.g. in an at-risk child
vaccinated against hepatitis B) or to confirm prior infection (e.g.
cytomegalovirus (CMV)). IgG antibody tends to persist after infection, whereas
IgM antibody reflects recent infection. This can be important in the newborn
period for distinguishing congenital infection (e.g. syphilis) from maternal
infection since IgG antibody readily crosses the placenta. Antibody responses
to infection are often described as ‘titres’. The titre is the reciprocal of
the highest dilution of the patient’s serum in which antibody was detected, e.g. a titre of 1024 means that
antibody was detected in a 1:1024 dilution of serum. The higher the titre, the
more the anti-body is present. The anti-streptolysin-O titre (ASOT) is
sometimes used as a marker of streptococcal infection in rheumatic fever.
Direct detection methods
Molecular biology techniques can now identify certain organisms, such as
viruses, that have traditionally been difficult to culture. These tests can
either use immunofluorescence, e.g. to identify respiratory syncytial virus
(RSV) in pharyngeal secretions in a child with bronchiolitis, or polymerase
chain reaction (PCR) to amplify bacterial or viral DNA using specific primers.
PCR methods are available for many important paediatric infectious agents,
including herpes simplex virus (HSV), Neisseria meningitidis groups B
and C, and HIV. They are particularly useful in confirming infection after
antibiotics have already been given and in detecting viral CNS infection.
Lumbar puncture and CSF analysis
Lumbar puncture is usually performed to diagnose or exclude meningitis.
It should not be performed if there is evidence of raised intracranial
pressure, if the child is haemodynamically unstable (e.g. septic shock) or if
there is a low platelet count or coagulopathy. A fine spinal needle with a
stylet is passed between the vertebral spines into the subdural space. A few
drops of CSF are collected for microbiological examination and for analysis of
protein and glucose concentrations. Examination of CSF includes microscopy and
culture and may also include other direct detection techniques (e.g. DNA
detection with PCR). Normal CSF is usually ‘crystal clear’. If it is cloudy,
this suggests infection or bleeding. Fresh blood that clears usually indicates
a traumatic tap, but a massive intracranial haemorrhage must be considered if
the CSF remains bloodstained. Old blood gives a yellow ‘xanthochromic’
appearance. A manometer can be used to easure the CSF pressure, though this is not routinely performed.
Although these are typical
CSF findings for the organisms indicated, partially treated infection and
infection with specific microorganisms may result in alternative profiles. For
example, meningitis caused by Listeria monocytogenes usually presents
with a CSF lymphocytosis.
Urinalysis
Fresh urine should be collected into a sterile container from a midstream
sample if possible. Urine bags applied over the genitalia may be used in
infants, but often become contaminated with perineal bacteria. Catheter
specimens and supra pubic aspirate are alternatives.
•
Observe the urine—is it cloudy (suggests
infection) or clear?
•
What is the colour?—pink or red suggests
haematuria from the lower urinary tract; brown (‘cola’-coloured) urine suggests
renal haematuria or haemolytic disease (haemoglobinuria).
•
Smell the urine for ketones and for the smell of
infection. Unusual-smelling urine may suggest an inborn error of metabolism.
•
Test the urine using commercial dipsticks. This
may reveal the following:
• protein—infection,
renal damage or nephrotic syndrome
• glucose—diabetes
• ketones—diabetic
ketoacidosis (DKA)
• white
cells or nitrites—suggestive of infection
•
These sticks are very sensitive to the presence
of blood and may detect haematuria even if the urine looks clear.
• Examine the urine under the microscope for white
cells, red cells, casts and the presence of organisms. A sample should also be
sent for culture. A pure growth of >105 colony-forming units of a
single organism and 50 white cells/mm3 confirms infection. Infection is extremely unlikely in the absence of
pyuria.
Immunology
Immunological in needed
to investigate a child with suspected immunodeficiency (severe, recurrent or
unusual infections—see Chapter 25) or with an autoimmune disorder such as
juvenile rheumatoid arthritis see Chapter 47) or systemic lupus erythematosus or renal disease.
Investigations should only be requested to confirm a clinical diagnosis,
or if indicated following a thorough history and examination. Sometimes, they
are performed to rule out more serious but less likely conditions. Blindly
performing investigations as a ‘fishing’ exercise in the hope of throwing up an
abnormality is usually counterproductive, often leading to increased anxiety
and further investigations when
unexpected results are obtained.
