Traceability and distribution of Neisseria meningitidis DNA in archived post mortem tissue samples from patients with systemic meningococcal disease

Berit Sletbakk Brusletto; Bernt Christian Hellerud; Else Marit Løberg; Ingeborg Løstegaard Goverud; Åshild Vege; Jens Petter Berg; Petter Brandtzaeg; Reidun Øvstebø

doi:10.1186/s12907-017-0049-9

Traceability and distribution of Neisseria meningitidis DNA in archived post mortem tissue samples from patients with systemic meningococcal disease

BMC Clin Pathol. 2017 Aug 16:17:10. doi: 10.1186/s12907-017-0049-9. eCollection 2017.

Authors

Berit Sletbakk Brusletto^{1

2}, Bernt Christian Hellerud³, Else Marit Løberg^{4

2}, Ingeborg Løstegaard Goverud^{4

2}, Åshild Vege^{5

2}, Jens Petter Berg^{1

2}, Petter Brandtzaeg^{1

2}, Reidun Øvstebø¹

Affiliations

¹ Blood Cell Research Group, Section for Research, Department of Medical Biochemistry, Oslo University Hospital HF, Ullevål Hospital, PO Box 4956 Nydalen, 0424 Oslo, Norway.
² Institute of Clinical Medicine, University of Oslo, Oslo, Norway.
³ Institute of Immunology, Oslo University Hospital, Oslo, Norway.
⁴ Department of Pathology, Oslo University Hospital, Oslo, Norway.
⁵ Section for Forensic Pediatric Pathology, Department of Forensic Sciences, Oslo University Hospital, Oslo, Norway.

Abstract

Background: The pathophysiology and outcome of meningococcal septic shock is closely associated with the plasma level of N. meningitidis lipopolysaccharides (LPS, endotoxin) and the circulating level of meningococcal DNA. The aim of the present study was to quantify the number of N. meningitidis in different formalin-fixed, paraffin-embedded (FFPE) tissue samples and fresh frozen (FF) tissue samples from patients with systemic meningococcal disease (SMD), to explore the distribution of N. meningitidis in the body.

Methods: DNA in FFPE and FF tissue samples from heart, lungs, liver, kidneys, spleen and brain from patients with meningococcal shock and controls (lethal pneumococcal infection) stored at variable times, were isolated. The bacterial load of N. meningitidis DNA was analyzed using quantitative real-time PCR (qPCR) and primers for the capsule transport A (ctrA) gene (1 copy per N. meningitidis DNA). The human beta-hemoglobin (HBB) gene was quantified to evaluate effect of the storage times (2-28 years) and storage method in archived tissue.

Results: N. meningitidis DNA was detected in FFPE and FF tissue samples from heart, lung, liver, kidney, and spleen in all patients with severe shock. In FFPE brain, N. meningitidis DNA was only detected in the patient with the highest concentration of LPS in the blood at admission to hospital. The highest levels of N. meningitidis DNA were found in heart tissue (median value 3.6 × 10⁷ copies N. meningitidis DNA/μg human DNA) and lung tissue (median value 3.1 × 10⁷ copies N. meningitidis DNA/μg human DNA) in all five patients. N. meningitidis DNA was not detectable in any of the tissue samples from two patients with clinical meningitis and the controls (pneumococcal infection). The quantity of HBB declined over time in FFPE tissue stored at room temperature, suggesting degradation of DNA.

Conclusions: High levels of N. meningitidis DNA were detected in the different tissue samples from meningococcal shock patients, particularly in the heart and lungs suggesting seeding and major proliferation of meningococci in these organs during the development of shock, probably contributing to the multiple organ failure. The age of archived tissue samples appear to have an impact on the amount of quantifiable N. meningitidis DNA.

Keywords: FFPE; Neisseria meningitidis; Quality of archived tissue; Systemic meningococcal disease; ctrA gene.