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Utility of Immunohistochemistry to Diagnose Cat Scratch Disease

P170

GC Caponetti , L Pantanowitz , LW Lamps , S Marconi , JM Havens and CN Otis .


1

Department of Pathology, Baystate Medical Center, Tufts University School of Medicine, Springfield, MA,
2
and Department of Pathology, University of Arkansas for Medical Sciences, Little Rock, AR.

ABSTRACT

METHODS

Background: Cat scratch disease (CSD) is largely due to infection with Bartonella henselae (BH).
Since BH is difficult to culture, the diagnosis of CSD relies mainly on clinical, serological and/or
histological evaluation. The latter requires stellate suppurative granulomas. Silver stains (e.g.
Warthin-Starry or Steiner) for bacteria are cumbersome to perform and hard to interpret.
Molecular testing is not readily available. A monoclonal antibody (mAB) to BH has become
commercially available. The aim of this study was to evaluate the utility of immunohistochemistry
(IHC) to diagnose CSD on routine surgical specimens.

27 archival formalin-fixed, paraffin-embedded (study) cases of CSD


lymphadenitis were investigated.
Cases were included if CSD was a clinical consideration and if suppurative
granulomas were present (Figure 1).

Design: 26 archival formalin-fixed, paraffin-embedded (study) cases of CSD lymphadenitis were


investigated. Cases were included if CSD was a clinical consideration and if suppurative granulomas
were present. Controls (n=11) included toxoplasmosis (2), sarcoidosis (2), Hodgkin lymphoma (1),
dermatopathic lymphadenopathy (1), Rosai-Dorfman disease (1) and non-specific reactive lymphoid
hyperplasia (4). 5 micron tissue sections were immunostained with BH mAB at 1:4000, 1:800 and
1:100 dilutions (clone H2A10, Biocare Medical, Concord, CA). PCR was performed following
deparaffinization on 50 micron lesional tissue sections. Primer pairs targeted a 153-bp fragment of
the 16S rRNA gene present in both BH and B. quintana.
Result: Five (19%) study cases were positive by IHC. Immunoreactive bacteria were mainly cocci
located predominantly within suppurative granulomas. Nine (35%) study cases were positive by
PCR, only one of which was also positive by IHC (at 1:100 dilution only). All controls were negative
for both PCR and IHC.
Conclusion: These data show that although the overall diagnostic sensitivity of these tests is low for
CSD, PCR is more sensitive than IHC using mAB for BH. IHC positive cases with negative PCR may
be attributed to sampling from non-representative lesional tissue. Therefore, we recommend
performing PCR on all IHC negative cases suspected of having CSD.
The authors mention references to commercial products in this abstract.

INTRODUCTION
Cat scratch disease (CSD) is largely due to infection with Bartonella
henselae.
Since Bartonella henselae is difficult to culture, the diagnosis of CSD relies
mainly on clinical, serological and/or histological evaluation. The latter
requires stellate suppurative granulomas.
Silver stains (e.g. Warthin-Starry or Steiner) for bacteria are cumbersome to
perform and hard to interpret.
Molecular testing is not readily available1.
Immunohistochemistry using a monoclonal antibody (mAB) to Bartonella
henselae has become commercially available2-4.

OBJECTIVE
The aim of this study was to evaluate the utility of immunohistochemistry
(IHC) to diagnose CSD on routine surgical specimens.

Figure 1: Lymph node showing suppurative granulomas seen in CSD


lymphadenitis (H&E) at low (A) and high magnification (B). Bacteria
identified within the areas of necrosis with silver stain.(C).
Warthin-Starry/Steiner stains were positive in 5/11 (45%) cases.
Controls (n=11) included toxoplasmosis (2), sarcoidosis (2), Hodgkin
lymphoma (1), dermatopathic lymphadenopathy (1), Rosai-Dorfman disease
(1) and non-specific reactive lymphoid hyperplasia (4).
5 micron tissue sections were immunostained with Bartonella henselae mAB
at 1:4000, 1:800 and 1:100 dilutions (clone H2A10, Biocare Medical, Concord,
CA).
PCR was performed following deparaffinization on 50 micron lesional tissue
sections. Primer pairs targeted a 153-bp fragment of the 16S rRNA gene
present in both Bartonella henselae and Bartonella quintana.

RESULTS
Five (19%) study cases were positive by IHC. Immunoreactive bacteria were
mainly cocci located predominantly within suppurative granulomas (Figure 2).
Nine (35%) study cases were positive by PCR, only one of which was also
positive by IHC (at 1:100 dilution only) (Table 1).
All controls were negative for both PCR and IHC.
Two cases were identified as positive by IHC at 1:4000, 1:800 and 1:100
dilutions, while were negative on PCR.
A third case was identified as positive by IHC only at a 1:100 dilution.
The third case was the only one that was positive both on IHC and PCR, and
that case was identified on IHC at a 1:100 dilution only.

Figure 2: Immunohistochemistry of CSD lymphadenitis with Bartonella


henselae mAB at 1-800 dilution at low (A) and at high magnification (B and
C).
IHC + (dilution)

IHC - (dilution)

PCR +

1(1:100)

8(1:100, 1:800, 1:4000)

PCR -

4(1:100, 1:800, 1:4000)

14(1:100, 1:800, 1:4000)

Table 1: Correlation of immunohistochemistry with PCR for Bartonella


henselae.

CONCLUSIONS
These data show that although the overall diagnostic sensitivity of
immunohistochemistry and PCR is low for CSD, PCR is more sensitive than
immunohistochemistry using mAB for Bartonella henselae.
Cases that were positive with mAB for Bartonella henselae but had a
negative PCR may be attributed to sampling from non-representative
lesional tissue.
We recommend performing PCR on all IHC negative cases suspected of
having CSD.

REFERENCES
1. Qian X, Jin L, Hayden RT, Macon WR, Lloyd RV. Diagnosis of cat scratch disease with Bartonella
henselae infection in formalin-fixed paraffin-embedded tissues by two different PCR assays. Diagn
Mol Pathol. 2005 Sep;14(3):146-51.
2. Cheuk W, Chan AK, Wong MC, Chan JK. Confirmation of diagnosis of cat scratch disease by
immunohistochemistry. Am J Surg Pathol. 2006 Feb;30(2):274-5.
3. Min KW, Reed JA, Welch DF, Slater LN. Morphologically variable bacilli of cat scratch disease are
identified by immunocytochemical labeling with antibodies to Rochalimaea henselae. Am J Clin
Pathol. 1994 May;101(5):607-10.
4. English CK, Wear DJ, Margileth AM, Lissner CR, Walsh GP. Cat-scratch disease. Isolation and
culture of the bacterial agent. JAMA. 1988 Mar 4;259(9):1347-52.

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