Transcript Document

BAMBINO GESU’
Children Hospital
HealthCare and
Research Institute
Rome - ITALY
[email protected]
Abstract
(revised)
nervous system Tuberculosis
Central
(CNS-TB) is the most severe form of TB
and frequently, is revealed as meningitis
TB infection. Difficulties and technical
hitches
to
obtain
enough
and
satisfactory samples helpful for a
microbiological TB diagnosis purpose
are well known. We try to understand if
and how the new Interefron-γ release
assay QuantiFeron TB Gold (Cellestis
Limited-Australia), is able to upgrade our
microbiological diagnostic protocol.
Methods: During the last three years
(2004-2007) we investigated at Bambino
Gesù Children Hospital in Rome, a total
of eight cases clinical suspected as TB
meningitis. For each one we applied the
microbiological
TB
investigation
protocol on Cerebrospinal fluid (CSF)
and respiratory specimens composed
by: Acid fast bacilli staining (AFB),
cultures growth in liquid and solid media
and nucleic acid amplification assay
(NAA). In adjunction we performed on
whole blood sample the QFT-G (liquid
antigen and In tube version).
Results : 4 out of 10 patients resulted
negative
for
microbiological
TB
investigation protocol and received, a
clinical diagnosis other than TB. About
the other four patients (median age 3 ys):
in three cases CSF resulted negative for
AFB and NAA was not performed
because insufficient; QFT-G was positive
in three case at the first evaluation (Time
0) and in two cases showed a conversion
of T-cell response: from Indeterminate to
Positive and from Negative to Positive at
a second assay performed into a new
collectected blood sample. All these four
TB cases were cultures confirmed.
Tuberculin Skin Test was performed in all
10 patients and resulted as Negative.
Discussion and Conclusion: As all
guidelines recommend, is mandatory to
use a correct microbiological approach
to avoid to lack TB diagnosis. Our on
field experience confirmed how is
difficult to obtain the right amount and
more than one sample of CNF in
children. On the other hand, the
introduction of a new not invasive
sample collection (peripheral whole
blood) and an easy to perform ELISA test
able to investigate a T cell specific
response
against
TB,
could
be
considerate a valid and rapid tool to
support
the
microbiological
TB
investigation protocol.
Presentation Number:U-008
Poster Board Number:0993
Background and Rationale of the Study
The clinical presentation of TB in children is extremely variable. It
depends by different factors as: the age, the immunocapability of the host
response and also the TB spread. In particular Tuberculous meningitis
(TBM) has high morbidity and mortality. Demonstration of tubercle bacilli
in cerebrospinal fluid (CSF), the only reliable method of diagnosis, is time
consuming and has a low yield. Also NAA commercial kits have low and
varying sensitivities, and therefore should not be used for excluding a
diagnosis of tuberculous meningitis. Moreover, cutaneous sensitivity to
Purified Protein antigen Derivative (TST) is not satisfied.
Results and Discussion
TB MENINGITIS
Culture
QFT- on blood
Patient
DA
EOA
GG
VD
Age
1y 2m
4y 3m
16y
1y 11m
1st
2nd
evaluation
evaluation
POS
IND
NEG
POS
7 days later
POS
14 days
later
POS
Specimen
Days
for
positivization
Identification
CSF
13 days
M. tuberculosis
Not Done
CSF
13 days
M. tuberculosis
Not Done
M. tuberculosis
CSF Not
Done
respiratory
samples
NEG
M. tuberculosis
CSF and
respiratory
samples
POS
CSF
CSF
14 days
11 days
NAA
Droplet infection
Primary pulmonary focus
bacteraemia spread seeding of
bacilli in meninges
thick exudate developed by Rich foci rupture into the
subarachnoid space. The inflammatory response gives:
• Adhesion (hydrocephalus)
• Vasculitis (internal carotid stenosis)
• Encephalitis (cerebral oedema)
Tab 1 Surveillance of Tuberculosis in Europe
In recent years, numerous authors studied possible biochemical markers
such as interferon gamma (IFN- ), to improve diagnostic efficiency.
The aims of our study have been:
to test if the new QuantiFeron TB Gold (QFT-G) is able to work in
suspected TB meningitis as rapid tool for TB diagnosis.
Population and Methods
OUR PATIENTS
QuantiFeronTB-Gold
From 2004 to 2007 we investigated
children admitted
at “Bambino Gesù” Children Hospital
(Rome, Italy) presenting with:
 Signs or symtoms of meningitis
compatible wich a TB suspected
INF- ELISA assay
Liquid and In Tube version
Step 1 samples collection
Step 2 ELISA assay
Step 3 Interpretation of INF-γ amount
Mycobacterial Laboratory Investigation Protocol

AFB
fluorescence
fluorescenza
Stain
 Solid
and liquid media
cultures
 Nucleid Acid
Amplification
BL
IM
1y4m
8m
IND
POS
POS
3 days later
CSF
12 days
M. tuberculosis
CSF and
respiratory
samples
NEG
Gastric
Aspirate
21 days
M. tuberculosis
NEG
Six children received the finaly diagnosis of TB meningitis
Five out six children had foreign origin (four from Est Europe,
one from Africa)
TST was not reactive (negative) in all children
QFT-G was positive during the first evaluation for three
children under 2 years of age
Computed Tomography images showed hydrocephalus and
basilar meningitis but with any evidence of tuberculoma
Our study was carried out prospectively in a clinical routinely
situation during the last three years.
We assessed whether the new Interferon-γ release assay
QuantiFERON-Gold could be used in practice in special setting (as
pediatric population) and in extra-pulmonary localization as TB
meningitis.
Potential limitations of our study should be:
400x
(Cobas Amplicore®- Roche)
the low number of cases may affect precision
we did not evaluate children HIV affected
Conclusions
Tuberculosis (TB) is a systemic infection caused by Mycobacterium tuberculosis. It can produce disease in any organ system by haematogenous
spread, specially in newborn and children. TB meningitis is the most severe form of TB and it may be suspected on the basis of clinical or
radiology experience. A correct diagnosis depends on the possibility to have a refererence mycobacteriology laboratory procedures for
appropriate investigation. Specimens for cultures often requiring invasive procedures and sophisticated laboratory techniques and they are Time
consuming. Our results reveal that: in case of suspected TB meningitis the QFT-G assay may offers rapid results expecially in children under
2 ys of age. In conclusion, in our experience QFT-G has revealed as a powerful tool to apply in a Mycobacterial laboratory investigation protocol
for diagnosis of suspected TB meningitis in children.
Acknowledgements
Silvia Gobbi and Eugenia Galeno laboratory technicians Microbiology Unit – Laboratory Department
Stefania Colafati MD for supporting in CT images and interpretation - Radiology Department