Galactomannan testing: lessons from the last decade

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Galactomannan testing lessons from the last
decade

• Claudio Viscoli
• Professor of Infectious Disease, University of
  Genova
• Chief, Division of Infectious Disease, San
  Martino University Hospital, Genova, Italy

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Galactomannan antigen detection Platelia
Aspergillus ELISA (Bio-Rad)
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Galactomannan antigenPlatelia Aspergillus
(Bio-Rad)

• Sensitivity highly variable (29-100)
• Specificity generally better (81-98)
• FDA approved
• Important tool in the diagnosis of aspergillosis
  (EORTC-MSG definitions of IA (Ascioglu 2002)
• May be positive before the occurrence of clinical
  and radiological signs/symptoms
• Two main strategies of use
• Serial collection of samples (2 or 3 times/week)
  in high risk patients
• Intensive testing in symptomatic patients
  (unexplained persistent fever unresponsive to
  broad spectrum antibiotics )

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Galactomannan antigenPlatelia Aspergillus
(Bio-Rad)

• Controversies
• Different cut-off used 0.5, 0.7, 1, 1.5
• Drawbacks
• False positive and false negative results
• Too low sensitivity according to some authors
  (Pinel 2003, Allan 2005)

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Galactomannan antigen CUT-OFF FOR POSITIVITY

• Test result as GM index sample OD/cut-off OD (1
  ng/ml )
• Index gt 1.5 in 2 consecutive samples (BIO-RAD)
• Index gt 1 (Verweij 1998 Maertens 2001
  Sulahian 2001 Ascioglu 2002)
• Index gt 0.7 (sensitivity24specificity-5.5 comp
  ared with BIO-RAD cut-off) (Herbrecht 2002)
• Index gt 0,5 (sensitivity 50?83,specificity
  100?73,7 compared with BIO-RAD cut-off
  (Marr 2004)

Single test Index gt 0.7
Static cut-off
(Maertens 2004)
Two consecutive test Index gt 0.5
Dynamic cut-off
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Galactomannan antigen
From 1998 to July 2009 24.093 Galactomannan
determinations with Platelia Aspergillus (ELISA)
(mean 2007 determinations/year min 332, max
4402)
We perform GM test in serum, BAL, sputum, CSF,
pleural fluid, tracheal aspirate fluid and
synovial fluid.
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Why we have false positive results?
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Aspergillus galactomannan False positive results

• Transient antigenemia (non invasive infections?)
• Cross reactivity with exoantigens
  (bacteria-fungi)
• Induction by cyclophosphamide (Hashiguchi et al.
  1994)
• Premature infants (83) (Siemann et al. 1998)
• Cotton swabs (Dalle et al. 2002)
• Absorption of galactomannan through a damaged
  intestinal mucosa (Letscher-Bru et al. 1998)
• During caspofungin therapy (Petraitiene et al.
  2002)
• Galactomannan in antibiotics (Ansorg et al. 1997
  Viscoli et al 2003)

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Fungal organism likely testing positive with the
Platelia test
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Routine use of the GM test at the BMT Unit in
Genova from Jan. 1999 to May 2003

• Total number of patients 420
• Total number of serum samples 4702
• Median samples per patient 7 (1-64)
• Median samples per month 85 (35-146)
• Median positivity rate per month
• Jan. 1999 - Jan. 2003 9 (0-18)
• Feb. 2003 - May 2003 24 (20-44)

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36 of patients and 28 of specimens were
positive
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Platelia Aspergillus Test results by
administration of Piperacillim-Tazobactam
Patient receiving piperacillin-tazobactam
Patient NOT receiving piperacillin-tazobactam
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89
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74
Pipera-tazo YES since at least 24 hrs
p lt 0,001
Viscoli et al ICAAC 2003 CID 2004
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Platelia Aspergillus teston piperacillin-tazobact
am

• six batches of Tazocin taken from the hospital
  pharmacy were tested
• two 4.5 g. vials per batch
• diluted with 100 ml NaCl 0.9
• five of six batches tested positive
• median GM index 4.7 (1.5-5.7)

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• Galactomannan antigen FALSE POSITIVE IN
  PEDIATRIC PATIENTS

• False positive GM test in 83 of premature
  infants
• (prolonged ICU and birth weight of 400-1320 g)
• (Siemann 1998)
• Passage of food-GM through damaged intestinal
  mucosa of BMT children (Letscher-Bru 1998)
• Neonates milk formula, false positive GM test
  (Gangneux 2002)
• Bifidobacterium sp. lipoteichoic acid (bacteria
  that heavily colonize neonatal gut) produces
  false positive GM test (Mennink-Kersten 2004)

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Clinical Microbiology and Infection, in press
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Why we have false negative results?

• Low prevalence of the disease
• Concomitant use of antifungals
• Little angioinvasion (HSCT)
• Presence of anti-aspergillus antibodies
• Low fungal burden
• Inappropriate cut-off
• Inappropriate use
• Testing
• Sampling
• Storage

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Pfeiffer et al., CID, 2006
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Antifungal therapy
Yes
No

• 0,5

• 1

• 1,5

• 1,5

• 1

• 0,5

(Marr 2005)
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Filtration and use of a larger volume of serum
Conventional method
Verwej 2005
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Galactomannan in other body fluids
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GM in CSF
(Klont RR, CID, 2004)
Cerebral aspergillosis 10-20 of all acses of
invasive aspergillosis

• Not validated
• Cut-off?

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Aspergillus galactomannan antigen detection in
cerebral aspergillosis
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Galactomannan as a surrogate marker of efficacy
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Galactomannan levels in serum and CSF samples
Sample / cut-off OD index
Days from BMT
(Machetti et al. 2000)
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Thank you for your attention
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Pfeiffer et al., CID, 2006
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Comparison of empirical and PCR-based
preemptive antifungal therapy in 408 allogeneic
stem cell transplant recipients

• PCR screening twice weekly during stay in
  hospital and once weekly after discharge until
  D100
• Antifungal therapy initiation
• PCR group in PCR patients with signs of
  infection and in patients with 2 consecutive PCR
  
• Empirical treatment group 5d of febrile
  neutropenia
• PCR based Empiric
• n 196 n 207
• Antifungal therapy 109 (56) 76 (37) (plt0.05)
• Proven invasive aspergilosis 11 16
• Reduction in early mortality (D30) in patients
  receiving PCR-based therapy but no difference in
  mortality at D100 and D180

• (Hebart et al. ASH 2004)