Poster Presentation Australian Society for Microbiology Annual Scientific Meeting 2013

Applicability of a Taqman PCR assay for the detection of Cryptococcus neoformans (#201)

Wafa Abdallah 1 , Francesca Azzato 1 , Norbert Ryan 1 , David E Leslie 1
  1. Victorian Infectious Disease Reference Laboratory, North Melbourne, VIC, Australia

Cryptococcus neoformans is an encapsulated yeast usually found in soil. Infection occurs predominantly in immunocompromised hosts, especially those with HIV infections. Initial infection commences with lung infection followed by haematogenous spread to the brain and meninges.
Methods for the diagnosis of Cryptococcus infection include culture, India ink staining of CSF and Cryptococcal antigen EIA or latex agglutination assays to detect capsular antigen (LAT). The Meridian (CALS) LAT is used in the Bacteriology laboratory at VIDRL for the diagnosis and subsequent monitoring of Cryptococcal infection. Although this assay has high specificity, problems may be encountered. A prozone phenomenon has been reported and it is now recognized that for HIV patients some isolates of C.neoformans may be poorly encapsulated. Additionally an antigen of Trichosporon beigelii has been recorded as cross-reacting in this test. Cryptococcal antigen titres are monitored long-term as a means of testing progress of disease post therapy. However variations in titre may be very difficult to interpret.
In an attempt to overcome these problems a Taqman PCR assay targeting the ITS1-ITS2 region overlapping the 5.8S rDNA gene of Cryptococcus neoformans (accession number JQ070099) was used. Screening of 20 CSF specimens positive by LAT gave only four positive PCR results. However these four specimens were collected at the time of initial diagnosis. Specimens taken post-treatment tested negative despite elevated antigen levels. Initial results demonstrate that this test may hold promise in determining active infection and monitoring treatment progress. Future development will include trialing of this assay for the screening of respiratory specimens from at risk patients.